Substituted heteroaryl benzofuran acids

ABSTRACT

The present invention relates generally to substituted heteroaryl benzofuran acids and methods of using them.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of prior U.S. patent applicationSer. No. 10/947,840 filed Sep. 23, 2004 which claims benefit of U.S.Provisional Application No. 60/506,012 filed Sep. 25, 2003, the entiredisclosure of which is incorporated herein by reference.

BACKGROUND

The present invention relates generally to substituted heteroarylbenzofuran acids and methods of using them.

The serine protease inhibitor PAI-1 is one of the primary inhibitors ofthe fibrinolytic system. The fibrinolytic system includes the proenzymeplasminogen, which is converted to the active enzyme, plasmin, by one oftwo tissue type plasminogen activators, t-PA or u-PA. PAI-1 is theprincipal physiological inhibitor of t-PA and u-PA. One of plasmin'smain responsibilities in the fibrinolytic system is to digest fibrin atthe site of vascular injury. The fibrinolytic system, however, is notonly responsible for the removal of fibrin from circulation but is alsoinvolved in several other biological processes including ovulation,embryogenesis, intima proliferation, angiogenesis, tumorigenesis, andatherosclerosis.

Elevated levels of PAI-1 have been associated with a variety of diseasesand conditions including those associated with impairment of thefibrinolytic system. For example, elevated levels of PAI-1 have beenimplicated in thrombotic diseases, e.g., diseases characterized byformation of a thrombus that obstructs vascular blood flow locally ordetaches and embolizes to occlude blood flow downstream. (Krishnamurti,Blood, 69, 798 (1987); Reilly, Arteriosclerosis and Thrombosis, 11, 1276(1991); Carmeliet, Journal of Clinical Investigation, 92, 2756 (1993),Rocha, Fibrinolysis, 8, 294, 1994; Aznar, Haemostasis 24, 243 (1994)).Antibody neutralization of PAI-1 activity resulted in promotion ofendogenous thrombolysis and reperfusion (Biemond, Circulation, 91, 1175(1995); Levi, Circulation 85, 305, (1992)). Elevated levels of PAI-1have also been implicated in diseases such as polycystic ovary syndrome(Nordt, Journal of clinical Endocrinology and Metabolism, 85, 4, 1563(2000)), bone loss induced by estrogen deficiency (Daci, Journal of Boneand Mineral Research, 15, 8, 1510 (2000)), cystic fibrosis, diabetes,chronic periodontitis, lymphomas, diseases associated with extracellularmatrix accumulation, malignancies and diseases associated withneoangiogenesis, inflammatory diseases, vascular damage associated withinfections, and diseases associated with increased uPA levels such asbreast and ovarian cancer.

In view of the foregoing, there exists a need for the identification ofinhibitors of PAI-1 activity and for methods of using the identifiedinhibitors to modulate PAI-1 expression or activity in a subject inorder to treat disorders associated with elevated PAI-1 levels.

SUMMARY

The present invention provides substituted heteroaryl benzofuran acidsand methods of using them. In certain embodiments, substitutedheteroaryl benzofuran acids of the invention include those of thefollowing formula:

wherein:

R, R₁, R₂, and R₃ are, independently, hydrogen, alkyl, cycloalkyl,alkanoyl, halo, hydroxy, aryl, perfluoroalkyl, alkoxy, amino,alkylamino, dialkylamino, or perfluoroalkoxy;

R₄ is hydrogen, alkyl, perfluoroalkyl, alkenyl, alkynyl, arylalkenyl,arylalkynyl, aryl, —C(═O)R₅, —C(═S)R₅, —CH₂R₅, or —CH(OH)R₅.

R₅ is hydrogen, C₁-C₈ alkyl, C₃-C₈ cycloalkyl, perfluoroalkyl, aryl,alkenyl, alkynyl, arylalkyl, arylalkenyl, arylalkynyl, alkylaryl,alkenylaryl, or alkynylaryl;

R₆ is hydrogen, alkyl, aryl, or arylalkyl;

x₁, x₂, x₃, x₄, x₅, x₆, x₇, and x₈ are, independently, carbon ornitrogen, wherein at least one of x₁-x₈ is a nitrogen atom;

n is an integer from 0-6; and

A is COOH or an acid mimic.

In certain embodiments when A is carboxylic acid, n is an integer from 1to 6.

The present invention also provides, inter alia, pharmaceuticallyacceptable salt or ester forms of formula 1.

The present invention further provides, inter alia, methods of usingsubstituted heteroaryl benzofuran acids. In one aspect of the presentinvention, a therapeutically effective amount of one or more substitutedheteroaryl benzofuran acids is administered to a subject in order totreat a PAI-1 related disorder, e.g., by inhibiting PAI-1 activity inthe subject. PAI-1 activity is associated with a number of diseases andconditions. For example, in one embodiment of the present invention,PAI-1 activity is associated with impairment of the fibrinolytic system.In other embodiments, PAI-1 activity is associated with thrombosis,e.g., venous thrombosis, arterial thrombosis, cerebral thrombosis, anddeep vein thrombosis, atrial fibrillation, pulmonary fibrosis,thromboembolic complications of surgery, cardiovascular disease, e.g.,myocardial ischemia, atherosclerotic plaque formation, chronicobstructive pulmonary disease, renal fibrosis, polycystic ovarysyndrome, Alzheimer's disease, or cancer.

DETAILED DESCRIPTION A. General Overview

The present invention provides compounds that inhibit PAI-1 activity,processes for preparing such compounds, pharmaceutical compositionscontaining such compounds, and methods for using such compounds inmedical therapies. The compounds have properties that are useful for thetreatment, including the prevention and inhibition, of a wide variety ofdiseases and disorders including those involving the production and/oraction of PAI-1. These include disorders resulting from impairment ofthe fibrinolytic system including, but not limited to, thrombosis,coronary heart disease, renal fibrosis, atherosclerotic plaqueformation, pulmonary disease, myocardial ischemia, atrial fibrillation,coagulation syndromes, thromboembolic complications of surgery,peripheral arterial occlusion and pulmonary fibrosis. Other disordersinclude, but are not limited to, polycystic ovary syndrome, Alzheimer'sdisease, and cancer.

The terms “alkyl” and “alkylene,” as used herein, whether used alone oras part of another group, refer to substituted or unsubstitutedaliphatic hydrocarbon chains, the difference being that alkyl groups aremonovalent (i.e., terminal) in nature whereas alkylene groups aredivalent and typically serve as linkers. Both include, but are notlimited to, straight and branched chains containing from 1 to about 12carbon atoms, preferably 1 to 6 carbon atoms, unless explicitlyspecified otherwise. For example, methyl, ethyl, propyl, isopropyl,butyl, i-butyl and t-butyl are encompassed by the term “alkyl.”Specifically included within the definition of “alkyl” are thosealiphatic hydrocarbon chains that are optionally substituted.Representative optional substituents include, but are not limited to,aryl, halogen, hydroxy, acyloxy, alkoxy, amino, amino substituted by oneor two alkyl groups of from 1 to 6 carbon atoms, aminoacyl, acylamino,thioalkoxy of from 1 to 6 carbon atoms, substituted thioalkoxy of from 1to 6 carbon atoms, and trihalomethyl. Preferred substituents includehalogen, trihalomethyl, alkyl of 1 to 6 carbon atoms, alkoxy of 1 to 6carbon atoms, and thioalkoxy of 1 to 6 carbon atoms. Preferably, alkyland alkylene groups are unsubstituted.

The carbon number as used in the definitions herein refers to carbonbackbone and carbon branching, but does not include carbon atoms of thesubstituents, such as alkoxy substitutions and the like.

The term “alkenyl”, as used herein, whether used alone or as part ofanother group, refers to a substituted or unsubstituted aliphatichydrocarbon chain and includes, but is not limited to, straight andbranched chains having 2 to 10 carbon atoms and containing at least onedouble bond. Preferably, the alkenyl moiety has 1 or 2 double bonds.Such alkenyl moieties can exist in the E or Z conformations and thecompounds of this invention include both conformations. Specificallyincluded within the definition of “alkenyl” are those aliphatichydrocarbon chains that are optionally substituted. Representativeoptional substituents include, but are not limited to, aryl, halogen,oxo (═O), hydroxy, acyloxy, alkoxy, amino, amino substituted by one ortwo alkyl groups of from 1 to 6 carbon atoms, aminoacyl, acylamino,thioalkoxy of from 1 to 6 carbon atoms, substituted thioalkoxy of from 1to 6 carbon atoms, and trihalomethyl. Preferred substituents includehalogen, trihalomethyl, alkyl of 1 to 6 carbon atoms, alkoxy of 1 to 6carbon atoms, and thioalkoxy of 1 to 6 carbon atoms. Heteroatoms, suchas O or S attached to an alkenyl should not be attached to a carbon atomthat is bonded to a double bond. Preferably, alkenyl groups areunsubstituted.

The term “alkynyl”, as used herein, whether used alone or as part ofanother group, refers to a substituted or unsubstituted aliphatichydrocarbon chain and includes, but is not limited to, straight andbranched chains having 2 to 10 carbon atoms and containing at least onetriple bond. Preferably, the alkynyl moiety has 3 to 6 carbon atoms. Incertain embodiments, the alkynyl can contain more than one triple bondand, in such cases, the alkynyl group must contain at least three carbonatoms. Specifically included within the definition of “alkynyl” arethose aliphatic hydrocarbon chains that are optionally substituted.Representative optional substituents include, but are not limited to,aryl, halogen, hydroxy, acyloxy, alkoxy, amino, amino substituted by oneor two alkyl groups of from 1 to 6 carbon atoms, aminoacyl, acylamino,thioalkoxy of from 1 to 6 carbon atoms, substituted thioalkoxy of from 1to 6 carbon atoms, and trihalomethyl. Heteroatoms, such as O or Sattached to an alkynyl should not be attached to the carbon that isbonded to a triple bond. Preferably, alkynyl groups are unsubstituted.

The term “cycloalkyl” as used herein, whether alone or as part ofanother group, refers to a substituted or unsubstituted alicyclichydrocarbon group having 3 to about 20 carbon atoms, preferably 3 to 8carbon atoms. Specifically included within the definition of“cycloalkyl” are those alicyclic hydrocarbon groups that are optionallysubstituted. Exemplary cycloalkyl groups include, but are not limitedto, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclooctyl,adamantyl, 2-[4-isopropyl-1-methyl-7-oxa-bicyclo[2.2.1]heptanyl], and2-[1,2,3,4-tetrahydro-naphthalenyl]. Representative optionalsubstituents include, but are not limited to, aryl, halogen, hydroxy,acyloxy, alkoxy, amino, amino substituted by one or two alkyl groups offrom 1 to 6 carbon atoms, aminoacyl, acylamino, thioalkoxy of from 1 to6 carbon atoms, substituted thioalkoxy of from 1 to 6 carbon atoms, andtrihalomethyl. Preferably, cycloalkyl groups are unsubstituted.

The term “aryl”, as used herein, whether used alone or as part ofanother group, is defined as a substituted or unsubstituted aromatichydrocarbon ring group having 5 to about 50 carbon atoms with form about6 to about 14 carbon atoms being preferred. The “aryl” group can have asingle ring or multiple condensed rings. The term “aryl” includes, butis not limited to phenyl, α-naphthyl, β-naphthyl, biphenyl, anthryl,tetrahydronaphthyl, fluorenyl, indanyl, biphenylenyl, and acenaphthenyl.Specifically included within the definition of “aryl” are those aromaticgroups that are optionally substituted. Accordingly, the aryl groupsdescribed herein refer to both unsubstituted or substituted aryl groups.For example, in representative embodiments of the present invention,the, “aryl” groups are optionally substituted with from 1 to 5substituents selected from the group consisting of acyloxy, hydroxy,acyl, alkyl of 1 to 6 carbon atoms, alkoxy of 1 to 6 carbon atoms,alkenyl of 2 to 6 carbon atoms, alkynyl of 2 to 6 carbon atoms, amino,amino substituted by one or two alkyl groups of from 1 to 6 carbonatoms, aminoacyl, acylamino, azido, cyano, halo, nitro, thioalkoxy offrom 1 to 6 carbon atoms, substituted thioalkoxy of from 1 to 6 carbonatoms, trihalomethyl, and aryl. Preferred substituents include halogen,trihalomethyl, alkyl of 1 to 6 carbon atoms, alkoxy of 1 to 6 carbonatoms, thioalkoxy of 1 to 6 carbon atoms, and aryl.

As used herein, the term “heteroaryl”, whether used alone or as part ofanother group, is defined as a substituted or unsubstituted aromaticheterocyclic ring system (monocyclic or bicyclic). Heteroaryl groups canhave, for example, from about 3 to about 50 carbon atoms (unlessexplicitly specified otherwise) with from about 4 to about 10 beingpreferred. In some embodiments, heteroaryl groups are aromaticheterocyclic rings systems having about 4 to about 14 ring atoms andcontaining carbon atoms and 1, 2, 3, or 4 heteroatoms selected fromoxygen, nitrogen or sulfur. Representative heteroaryl groups are furan,thiophene, indole, azaindole, oxazole, thiazole, isoxazole, isothiazole,imidazole, N-methylimidazole, pyridine, pyrimidine, pyrazine, pyrrole,N-methylpyrrole, pyrazole, N-methylpyrazole, 1,3,4-oxadiazole,1,2,4-triazole, 1-methyl-1,2,4-triazole, 1H-tetrazole,1-methyltetrazole, benzoxazole, benzothiazole, benzofuran,benzisoxazole, benzimidazole, N-methylbenzimidazole, azabenzimidazole,indazole, quinazoline, quinoline, and isoquinoline. Bicyclic aromaticheteroaryl groups include phenyl, pyridine, pyrimidine or pyridizinerings that are (a) fused to a 6-membered aromatic (unsaturated)heterocyclic ring having one nitrogen atom; (b) fused to a 5- or6-membered aromatic (unsaturated) heterocyclic ring having two nitrogenatoms; (c) fused to a 5-membered aromatic (unsaturated) heterocyclicring having one nitrogen atom together with either one oxygen or onesulfur atom; or (d) fused to a 5-membered aromatic (unsaturated)heterocyclic ring having one heteroatom selected from O, N or S.Specifically included within the definition of “heteroaryl” are thosearomatic heterocyclic rings that are substituted, for example with 1 to5 substituents selected from the group consisting of acyloxy, hydroxy,acyl, alkyl of 1 to 6 carbon atoms, alkoxy of 1 to 6 carbon atoms,alkenyl of 2 to 6 carbon atoms, alkynyl of 2 to 6 carbon atoms, amino,amino substituted by one or two alkyl groups of from 1 to 6 carbonatoms, aminoacyl, acylamino, azido, cyano, halo, nitro, thioalkoxy offrom 1 to 6 carbon atoms, substituted thioalkoxy of from 1 to 6 carbonatoms, and trihalomethyl.

The term “alkoxy” as used herein, refers to the group R_(a)—O— whereinR_(a) is an alkyl group as defined herein. Specifically included withinthe definition of “alkoxy” are those alkoxy groups that are optionallysubstituted.

The term “perfluoroalkyl”, as used herein, whether used alone or as partof another group, refers to a saturated aliphatic hydrocarbon having 1to about 10, preferably 1 to about 6 carbon atoms and two or morefluorine atoms and includes, but is not limited to, straight or branchedchains, such as —CF₃, —CH₂CF₃, —CF₂CF₃ and —CH(CF₃)₂.

The term “perfluoroalkoxy”, as used herein refers to the group R_(a)—O—wherein R_(a) is a perfluoroalkyl group as defined herein.

The term “alkylaryl”, as used herein, whether used alone or as part ofanother group, refers to the group —R_(a)—R_(b), where R_(a) is an arylgroup as defined above, substituted by R_(b), an alkyl group, as definedabove. Alkylaryl groups can be optionally substituted.

The term “arylalkyl” or “aralkyl” refers to the group —R_(a)—R_(b),where R_(a) is an alkylene group as defined above, substituted by R_(b),an aryl group, as defined above. Preferably the alkyl group has from 1to 6 carbon atoms. Examples of arylalkyl moieties include, but are notlimited to, benzyl, 1-phenylethyl, 2-phenylethyl, 3-phenylpropyl,2-phenylpropyl and the like. Arylalkyl groups can be optionallysubstituted.

The term “arylalkenyl”, as used herein, refers to the group—R_(a)—R_(b), where R_(a) is an alkene group, as defined above,substituted by R_(b), an aryl group, as defined above. Arylalkenylgroups can be optionally substituted.

The term “arylalkynyl” as used herein, refers to the group —R_(a)—R_(b),where R_(a) is an alkyne group, as defined above, substituted by R_(b),an aryl group, as defined above. Arylalkynyl groups can be optionallysubstituted.

The term “alkylaryl”, as used herein, refers to the group —R_(b)—R_(a),where R_(b) is an aryl group, as defined above, substituted by R_(a), analkyl group, as defined above. Alkylaryl groups can be optionallysubstituted.

The term “alkenylaryl”, as used herein, refers to the group—R_(b)—R_(a), where R_(b) is an aryl group, as defined above,substituted by R_(a), an alkenyl group, as defined above. Alkenylarylgroups can be optionally substituted.

The term “alkynylaryl” as used herein, refers to the group —R_(b)—R_(a)where R_(b) is an aryl group as defined above, substituted by R_(a), analkynyl group, as defined above. Alkynylaryl groups can be optionallysubstituted.

The term “alkanoyl” as used herein, refers to the group —C(═O)-alkylwherein alkyl is defined as above. Exemplary alkanoyl groups include,but are not limited to, acetyl (ethanoyl), n-propanoyl, n-butanoyl,2-methylpropanoyl, n-pentanoyl, 2-methylbutanoyl, 3-methylbutanoyl,2,2-dimethylpropanoyl, heptanoyl, and decanoyl. The alkyl moieties ofalkanoyl groups can be optionally substituted.

The term “aroyl” refers to a carbonyl —C(═O)-aryl group wherein aryl isas previously defined. The aryl moieties of aroyl groups can beoptionally substituted. Exemplary aroyl groups include, but are notlimited to, benzoyl.

The term “alkylamino” refers to the group R_(c)—NH—, wherein R_(c) is analkyl group as defined above preferably having from 1 to 6 carbon atoms.

The term “dialkylamino” refers to the group —N(C₁-C₆ alkyl)₂.

The term “halogen” or “halo” refers to chlorine, bromine, fluorine, andiodine.

The term “treating” or “treatment” refers to any indicia of success inamelioration of an injury, pathology, or condition, including anyobjective or subjective parameter such as abatement; remission;diminishing of symptoms or making the injury, pathology, or conditionmore tolerable to the patient; slowing in the rate of degeneration ordecline; making the final point of degeneration less debilitating; orimproving a subject's physical or mental well-being. The treatment oramelioration of symptoms can be based on objective or subjectiveparameters; including the results of a physical examination,neurological examination, and/or psychiatric evaluation. “Treating” or“treatment of a PAI-1 related disorder” includes preventing the onset ofsymptoms in a subject that may be predisposed to a PAI-1 relateddisorder but does not yet experience or exhibit symptoms of the disorder(prophylactic treatment), inhibiting the symptoms of the disorder(slowing or arresting its development), providing relief from thesymptoms or side-effects of the disorder (including palliativetreatment), and/or relieving the symptoms of the disorder (causingregression). Accordingly, the term “treating” includes theadministration of the compounds or agents of the present invention to asubject to prevent or delay, to alleviate, or to arrest or inhibitdevelopment of the symptoms or conditions associated with PAI-1 relateddisorders, e.g., tumor growth associated with cancer. A skilled medicalpractitioner will know how to use standard methods to identify a subjectin need of treatment, e.g., by examining the patient and determiningwhether the patient is suffering from a disease known to be associatedwith elevated PAI-1 levels or activity or by assaying for PAI-1 levelsin blood plasma or tissue of the individual suspected of suffering froma PAI-1 related disease and comparing PAI-1 levels in the blood plasmaor tissue of the individual suspected of suffering from a PAI-1 relateddisease to PAI-1 levels in the blood plasma or tissue of a healthyindividual. Increased PAI-1 levels are indicative of disease.Accordingly, the present invention provides, inter alia, methods ofadministering a compound of the present invention to a subject anddetermining levels of PAI-1 in the subject. The level of PAI-1 in thesubject can be determined before and/or after administration of thecompound.

In healthy individuals, PAI-1 is found at low levels in the plasma (forexample from about 5-26 ng/mL), but it is elevated in many PAI-1 relateddisorders, including, for example, atherosclerosis (Schneiderman J. et.al, Proc Natl Acad Sci 89: 6998-7002, 1992) deep vein thrombosis(Juhan-Vague I, et. al, Thromb Haemost 57: 67-72, 1987), and non-insulindependent diabetes mellitus (Juhan-Vague I, et. al, Thromb Haemost 78:565-660, 1997). PAI-1 stabilizes both arterial and venous thrombi,contributing respectively to coronary arterial occlusion inpost-myocardial infarction (Hamsten A, et. al. Lancet 2:3-9, 1987), andvenous thrombosis following post-operative recovery from orthopedicsurgery. (Siemens H J, et. al, J Clin Anesthesia 11: 622-629, 1999).Plasma PAI-1 is also elevated, for example, in postmenopausal women, andhas been proposed to contribute to the increased incidence ofcardiovascular disease in this population (Koh K et. al, N Engl J Med336: 683-690, 1997).

The term “PAI-1 related disorder or disease” refers to any disease orcondition that is associated with increased or enhanced expression oractivity of PAI-1 or increased or enhanced expression or activity of agene encoding PAI-1. Examples of such increased activity or expressioncan include one or more of the following: activity of the protein orexpression of the gene encoding the protein is increased above the levelof that in normal subjects; activity of the protein or expression of thegene encoding the protein is in an organ, tissue or cell where it is notnormally detected in normal subjects (i.e. spatial distribution of theprotein or expression of the gene encoding the protein is altered);activity of the protein or expression of the gene encoding the proteinis increased when activity of the protein or expression of the geneencoding the protein is present in an organ, tissue or cell for a longerperiod than in a normal subjects (i.e., duration of activity of theprotein or expression of the gene encoding the protein is increased). Anormal or healthy subject is a subject not suffering from a PAI-1related disorder or disease. In some embodiments of the presentinvention, the PAI-1 related disorder is not associated withhyperglycemia. A PAI-1 related disorder that is not associated withhyperglycemia is one, for example, that is not caused by elevated levelsof glucose in the blood.

The term “pharmaceutically acceptable excipient” means an excipient thatis useful in preparing a pharmaceutical composition that is generallysafe, non-toxic, and desirable, and includes excipients that areacceptable for veterinary use as well as for human pharmaceutical use.Such excipients can be solid, liquid, semisolid, or, in the case of anaerosol composition, gaseous.

“Pharmaceutically acceptable salts and esters” refers to salts andesters that are pharmaceutically acceptable and have the desiredpharmacological properties. Such salts include, for example, salts thatcan be formed where acidic protons present in the compounds are capableof reacting with inorganic or organic bases. Suitable inorganic saltsinclude, for example, those formed with the alkali metals or alkalineearth metals, e.g. sodium and potassium, magnesium, calcium, andaluminum. Suitable organic salts include, for example, those formed withorganic bases such as the amine bases, e.g. ethanolamine,diethanolamine, triethanolamine, trimethamine, N methylglucamine, andthe like. Pharmaceutically acceptable salts can also include acidaddition salts formed from the reaction of basic moieties, such asamines, in the parent compound with inorganic acids (e.g. hydrochloricand hydrobromic acids) and organic acids (e.g. acetic acid, citric acid,maleic acid, and the alkane- and arene-sulfonic acids such asmethanesulfonic acid and benzenesulfonic acid). Pharmaceuticallyacceptable esters include esters formed from carboxy, sulfonyloxy, andphosphonoxy groups present in the compounds, e.g. C₁₋₆ alkyl esters.When there are two acidic groups present, a pharmaceutically acceptablesalt or ester can be a mono-acid-mono-salt or ester or a di-salt orester; and similarly where there are more than two acidic groupspresent, some or all of such groups can be salified or esterified.Compounds named in this invention can be present in unsalified orunesterified form, or in salified and/or esterified form, and the namingof such compounds is intended to include both the original (unsalifiedand unesterified) compound and its pharmaceutically acceptable salts andesters. Also, certain compounds named in this invention can be presentin more than one stereoisomeric form, and the naming of such compoundsis intended to include all single stereoisomers and all mixtures(whether racemic or otherwise) of such stereoisomers.

“Inhibitors,” “activators,” and “modulators” of expression or ofactivity are used to refer to inhibitory, activating, or modulatingmolecules, respectively, identified using in vitro and in vivo assaysfor expression or activity. Inhibitors of the present invention arecompositions that, inhibit expression of PAI-1 or bind to, partially ortotally block stimulation, decrease, prevent, delay activation,inactivate, desensitize, or down regulate the activity of PAI-1. Samplesor assays comprising PAI-1 can be treated with a composition of thepresent invention and compared to control samples without a compositionof the present invention. Control samples (untreated with compositionsof the present invention) can be assigned a relative activity value of100%. In certain embodiments, inhibition of PAI-1 is achieved when theactivity value relative to the control is about 80% or less, optionally50% or 25, 10%, 5% or 1%.

The terms “pharmaceutically acceptable”, “physiologically tolerable” andgrammatical variations thereof, as they refer to compositions, carriers,diluents and reagents, are used interchangeably and represent that thematerials are capable of administration to or upon a human without theproduction of undesirable physiological effects such as nausea,dizziness, gastric upset and the like which would be to a degree thatwould prohibit administration of the compound.

A “therapeutically effective amount” or “pharmaceutically effectiveamount” means the amount that, when administered to a subject, produceseffects for which it is administered. For example, a “therapeuticallyeffective amount,” when administered to a subject to inhibit PAI-1activity, is sufficient to inhibit PAI-1 activity. A “therapeuticallyeffective amount,” when administered to a subject for treating adisease, is sufficient to effect treatment for that disease.

Except when noted, the terms “subject” or “patient” are usedinterchangeably and refer to mammals such as human patients andnon-human primates, as well as experimental animals such as rabbits,rats, and mice, and other animals. Accordingly, the term “subject” or“patient” as used herein means any mammalian patient or subject to whichthe compounds of the invention can be administered. In an exemplaryembodiment of the present invention, to identify subject patients fortreatment according to the methods of the invention, accepted screeningmethods are employed to determine risk factors associated with atargeted or suspected disease or condition or to determine the status ofan existing disease or condition in a subject. These screening methodsinclude, for example, conventional work-ups to determine risk factorsthat may be associated with the targeted or suspected disease orcondition. These and other routine methods allow the clinician to selectpatients in need of therapy using the methods and formulations of thepresent invention. In some embodiments of the present invention, thesubject to be treated with the methods of the present invention does nothave hyperglycemia and/or a disease that has been caused byhyperglycemia. Methods of determining whether a subject hashyperglycemia are known in the art and include, for example, performinga glucose test that measures the level of glucose in the blood. Twoexemplary tests that can be used to measure the presence of excesslevels of glucose in the blood include a test that measures the amountof glucose in the blood after an overnight fast and a test that measuresthe body's ability to process excess sugar presented after drinking ahigh glucose test. Typically a subject having a fasting sugar level(sugar level after an overnight fast) of about 64 to about 110 mg/dldoes not have hyperglycemia whereas as person having a fasting sugarlevel of greater than 110 mg/dl has elevated blood sugar levels. A valueabove about 140 mg/dl on at least two occasions typically signifies thatthe subject has diabetes. In some embodiments, the subject to be treatedwith the methods of the present invention is not otherwise in need oftreatment with a leukotriene antagonist.

When any variable occurs more than one time in any constituent or in anyformula, its definition in each occurrence is independent of itsdefinition at every other occurrence. Combinations of substituentsand/or variables are permissible only if such combinations result instable compounds.

B. Substituted Heteroaryl Benzofuran Acids

The present invention provides substituted heteroaryl benzofuran acids.Such compounds are preferably administered to inhibit PAI-1 expressionor activity in a subject and, ultimately, to treat diseases orconditions including those associated with increased PAI-1 activity in asubject, e.g., a PAI-1 related disorder.

Substituted heteroaryl benzofuran acids include those of the followingformula:

wherein:

R, R₁, R₂, and R₃ are, independently, hydrogen, alkyl, cycloalkyl,alkanoyl, halo, hydroxy, aryl, perfluoroalkyl, alkoxy, amino,alkylamino, dialkylamino, or perfluoroalkoxy;

R₄ is hydrogen, alkyl, perfluoroalkyl, alkenyl, alkynyl, arylalkenyl,arylalkynyl, aryl, —C(═O)R₅, —C(—S)R₅, —CH₂R₅, or —CH(OH)R₅.

R₅ is hydrogen, C₁-C₈ alkyl, C₃-C₈ cycloalkyl, perfluoroalkyl, aryl,alkenyl, alkynyl, arylalkyl, arylalkenyl, arylalkynyl, alkylaryl,alkenylaryl, or alkynylaryl;

R₆ is hydrogen, alkyl, aryl, or arylalkyl;

x₁, x₂, x₃, x₄, x₅, x₆, x₇, and x₈ are, independently, carbon ornitrogen, wherein at least one of x₁-x₈ is a nitrogen atom;

n is an integer from 0-6, preferably 1-6; and

A is COOH or an acid mimic.

Compounds of the present invention also include prodrugs, stereoisomers,or pharmaceutically acceptable salts or ester forms of Formula 1.

In some exemplary compounds of the present invention, R and R₁ arehydrogen and R₄ is hydrogen, alkyl, or aryl. In some preferredembodiments, R and R₁ are hydrogen and R₄ is hydrogen, C₅ alkyl, orphenyl. In such embodiments, R₂, R₃, R₅, R₆, n, A, and x₁, x₂, x₃, x₄,x₅, x₆, x₇, and x₈ are as defined herein for compounds of Formula 1.

In some exemplary compounds of Formula 1, R₂ and R₃ are, independently,hydrogen or perfluoroalkyl. In certain preferred embodiments, R₂ and R₃are, independently, hydrogen or CF₃. In such embodiments, R, R₁, R₄, R₅,n, A, and x₁, x₂, x₃, x₄, x₅, x₆, x₇, and x₈ are as defined herein forcompounds of Formula 1.

A can be COOH or an acid mimic. In some preferred embodiments ofcompounds of Formula 1, A is COOH or tetrazole. In such embodiments, R,R₁, R₂, R₃, R₄, R₅ R₆, n, and x₁, x₂, x₃, x₄, x₅, x₆, x₇, and x₈ are asdefined herein for compounds of Formula 1.

In certain preferred compounds of the present invention, R₆ is hydrogen.In such embodiments, R, R₁, R₂, R₃, R₄, R₅, n, A, and x₁, x₂, x₃, x₄,x₅, x₆, x₇, and x₈ are as defined herein for compounds of Formula 1.

In an exemplary embodiment of compounds of Formula 1, R₄ isperfluoroalkyl, alkenyl, arylalkenyl, aryl, —C(═O)R₅, —C(═S)R₅, —CH₃R₅,or —CH₂(OH)R₅ and R, R₁, R₂, R₃, R₅ R₆, n, A, and x₁, x₂, x₃, x₄, x₅,x₆, x₇, and x₈ are as defined herein for compounds of Formula 1.

In other embodiments, substituted heteroaryl benzofuran acids of theinvention include those of the following formula:

wherein:

R, R₁, R₂, and R₃ are, independently, hydrogen, alkyl, cycloalkyl,alkanoyl, halo, hydroxy, aryl, perfluoroalkyl, alkoxy, amino,alkylamino, dialkylamino, or perfluoroalkoxy;

R₄ is hydrogen, alkyl, perfluoroalkyl, alkenyl, alkynyl, arylalkenyl,arylalkynyl, aryl, —C(═O)R₅, —C(═S)R₅, —CH₂R₅, or —CH(OH)R₅.

R₅ is hydrogen, C₁-C₈ alkyl, C₃-C₈ cycloalkyl, perfluoroalkyl, aryl,alkenyl, alkynyl, arylalkyl, arylalkenyl, arylalkynyl, alkylaryl,alkenylaryl, or alkynylaryl;

R₆ is hydrogen, alkyl, aryl, or arylalkyl;

x₁, x₂, x₃, x₄, x₅, x₆, x₇, and x₈ are, independently, carbon ornitrogen, wherein at least one of x₁-x₈ is a nitrogen atom; and

A is an acid mimic.

Compounds of the present invention also include prodrugs, stereoisomers,or pharmaceutically acceptable salts or ester forms of Formula 2.

In certain exemplary compounds of Formula 2, R and R₁ are hydrogen andR₄ is hydrogen, alkyl, or aryl. In some preferred embodiments, R and R₁are hydrogen and R₄ is hydrogen, C₅, or phenyl. In such embodiments, R₂,R₃, R₅, R₆, A, and x₁, x₂, x₃, x₄, x₅, x₆, x₇, and x₈ are as definedherein for compounds of Formula 2.

In some exemplary compounds of Formula 2, R₂ and R₃ are, independently,hydrogen or perfluoroalkyl. In certain preferred embodiments, R₂ and R₃are, independently, hydrogen or CF₃. In such embodiments, R, R₁, R₄, R₅,R₆, A, and x₁, x₂, x₃, x₄, x₅, x₆, x₇, and x₈ are as defined herein forcompounds of Formula 2.

In some preferred embodiments of compounds of Formula 2, A is tetrazole.In such embodiments, R, R₁, R₂, R₃, R₄, R₅, R₆, and x₁, x₂, x₃, x₄, x₅,x₆, x₇, and x₈ are as defined herein for compounds of Formula 2.

In an exemplary embodiment of compounds of Formula 2, R₄ isperfluoroalkyl, alkenyl, arylalkenyl, aryl, —C(═O)R₅, —C(═S)R₅, —CH₃R₅,or —CH₂(OH)R₅ and R, R₁, R₂, R₃, R₅ R₆, n, A, and x₁, x₂, x₃, x₄, x₅,x₆, x₇, and x₈ are as defined herein for compounds of Formula 2.

Exemplary substituted heteroaryl benzofuran acids of the presentinvention include, but are not limited to,6-(1-Benzofuran-2-yl)-2-(trifluoromethyl)quinolin-4-yl1H-tetraazol-5-ylmethyl ether or a pharmaceutically acceptable salt orester form thereof,6-(3-Pentyl-1-benzofuran-2-yl)-4-(1H-tetraazol-5-ylmethoxy)-2-(trifluoromethyl)-quinolineor a pharmaceutically acceptable salt or ester form thereof,5-(1-Benzofuran-2-yl)-8-(1H-tetraazol-5-ylmethoxy)quinoline or apharmaceutically acceptable salt or ester form thereof,5-(3-Phenyl-1-benzofuran-2-yl)-8-quinolinyl 1H-tetraazol-5-ylmethylether or a pharmaceutically acceptable salt or ester form thereof, and5-(3-Pentyl-1-benzofuran-2-yl)-8-(1H-tetraazol-5-ylmethoxy)quinoline ora pharmaceutically acceptable salt or ester form thereof.

The present invention also provides compositions comprising substitutedheteroaryl benzofuran acids, including those compounds of formulas 1 and2 or a stereoisomer or pharmaceutically acceptable salt thereof, and oneor more pharmaceutically acceptable carriers, excipients, or diluents.Such compositions include pharmaceutical compositions for treating orcontrolling disease states or conditions associated with increased PAI-1activity. In certain embodiments, the compositions comprise mixtures ofone or more substituted heteroaryl benzofuran acids.

Certain of the compounds of formulas 1 and 2 contain stereogenic carbonatoms or other chiral elements and thus give rise to stereoisomers,including enantiomers and diastereomers. The present invention includesall of the stereoisomers of formulas 1 and 2, as well as mixtures of thestereoisomers. Throughout this application, the name of the product,where the absolute configuration of an asymmetric center is notindicated, is intended to embrace the individual stereoisomers as wellas mixtures of stereoisomers. When it is necessary to distinguish theenantiomers from one another and from the racemate, the sign of theoptical rotation [(+), (−) and (±)] is utilized. Furthermore, throughoutthis application, the designations R* and S* are used to indicaterelative stereochemistry, employing the Chemical Abstracts conventionwhich automatically assigns R* to the lowest numbered asymmetric center.

Where an enantiomer is preferred, it can, in some embodiments, beprovided substantially free of the corresponding enantiomer. Thus, anenantiomer substantially free of the corresponding enantiomer refers toa compound that is isolated or separated via separation techniques orprepared free of the corresponding enantiomer. “Substantially free,” asused herein, means that the compound is made up of a significantlygreater proportion of one enantiomer. In preferred embodiments, thecompound is made up of at least about 90% by weight of a preferredenantiomer. In other embodiments of the invention, the compound is madeup of at least about 99% by weight of a preferred enantiomer. Preferredenantiomers can be isolated from racemic mixtures by any method known tothose skilled in the art, including high performance liquidchromatography (HPLC) and the formation and crystallization of chiralsalts, or preferred enantiomers can be prepared by methods describedherein. Methods for the preparation of preferred enantiomers aredescribed, for example, in Jacques, et al., Enantiomers, Racemates andResolutions (Wiley Interscience, New York, 1981); Wilen, S. H., et al.,Tetrahedron 33:2725 (1977); Eliel, E. L. Stereochemistry of CarbonCompounds (McGraw-Hill, NY, 1962); and Wilen, S. H. Tables of ResolvingAgents and Optical Resolutions p. 268 (E. L. Eliel, Ed., Univ. of NotreDame Press, Notre Dame, Ind. 1972).

Exemplary salt forms of the compounds herein include, but are notlimited to, sodium salts and potassium salts. Other exemplary salt formsof these compounds include, but are not limited to, those formed withpharmaceutically acceptable inorganic and organic bases known in theart. Salt forms prepared using inorganic bases include hydroxides,carbonates or bicarbonates of the therapeutically acceptable alkalimetals or alkaline earth metals, such as sodium potassium, magnesium,calcium and the like. Acceptable organic bases include amines, such asbenzylamine, mono-, di- and trialkylamines, preferably those havingalkyl groups of from 1 to 6 carbon atoms, more preferably 1 to 3 carbonatoms, such as methylamine, dimethylamine, trimethylamine, ethylamine,diethylamine, triethylamine, mono-, di-, and triethanolamine. Exemplarysalts also include alkylene diamines containing up to 6 carbon atoms,such as hexamethylenediamine; cyclic saturated or unsaturated basescontaining up to 6 carbon atoms, including pyrrolidine, piperidine,morpholine, piperazine and their N-alkyl and N-hydroxyalkyl derivatives,such as N-methyl-morpholine and N-(2-hyroxyethyl)-piperidine, orpyridine. Quaternary salts can also be formed, such as tetralkyl forms,such as tetramethyl forms, alkyl-alkanol forms, such asmethyl-triethanol or trimethyl-monoethanol forms, and cyclic ammoniumsalt forms, such as N-methylpyridinium,N-methyl-N-(2-hydroxyethyl)-morpholinium, N,N-di-methylmorpholinium,N-methyl-N-(2-hydroxyethyl)-morpholinium, or N,N-dimethyl-piperidiniumsalt forms. These salt forms can be prepared using the acidiccompound(s) of Formulas 1 and 2 and procedures known in the art.

Exemplary ester forms of the compounds of this invention include, butare not limited to, straight chain alkyl esters having from 1 to 6carbon atoms or branched chain alkyl groups containing 1 to 6 carbonatoms, including methyl, ethyl, propyl, butyl, 2-methylpropyl and1,1-dimethylethyl esters, cycloalkyl esters, alkylaryl esters, benzylesters, and the like. Other exemplary esters include, but are notlimited to, those of the formula —COOR₁₀ wherein R₁₀ is selected fromthe formula:

wherein R₁₁, R₁₂, R₁₃, R₁₄ are independently selected from hydrogen,alkyl of from 1 to 10 carbon atoms, aryl of 6 to 12 carbon atoms,arylalkyl of from 6 to 12 carbon atoms; heteroaryl or alkylheteroarylwherein the heteroaryl ring is bound by an alkyl chain of from 1 to 6carbon atoms.

Acids and acid mimics, according to the invention, are defined as protonor hydrogen donating groups. Exemplary acid mimics or mimetics of thepresent invention include pharmaceutically useful carboxylic acids andacid mimics or mimetics known in the art, such as those described in R.Silverman, The Organic Chemistry of Drug Design and Drug Action,Academic Press (1992) and others. Exemplary acid mimics or mimeticsinclude tetrazole, tetronic acid, acyl tetronic acid, and groups havingthe formula:

wherein R₁₅ is C₁-C₆ alkyl, C₂-C₆ alkenyl, C₃-C₆ cycloalkyl, —CH₂—(C₃-C₆cycloalkyl), C₃-C₆ cycloalkenyl, —CH₂—(C₃-C₆ cycloalkenyl), optionallysubstituted aryl or heteroaryl groups or optionally substitutedaryl(C₁-C₆)alkyl or heteroaryl(C₁-C₆)alkyl, with the aryl and heteroarylgroups as defined herein.

Preferred compounds of the present invention inhibit PAI-1 activity.Accordingly, the compounds can be used for the treatment, includingprevention, inhibition, and/or amelioration of PAI-1 related disordersin a subject, including, for example, in the treatment of noninsulindependent diabetes mellitus, in the treatment of cardiovascular disease,and in the treatment of thrombotic events associated with coronaryartery and cerebrovascular disease. Using the methods of the presentinvention, a skilled medical practitioner will know how to administersubstituted heteroaryl benzofuran acids, including those represented byformulas 1 and 2, to a subject suffering from any of the diseasesassociated with increased PAI-1 activity or expression, e.g., diabetesor cardiovascular disease, in order to effect treatment for thatdisease.

In one exemplary embodiment, substituted heteroaryl benzofuran acids areadministered to a subject in order to treat disease processes involvingthrombotic and prothrombotic states which include, but are not limitedto, formation of atherosclerotic plaques, venous and arterialthrombosis, myocardial ischemia, atrial fibrillation, deep veinthrombosis, coagulation syndromes, pulmonary thrombosis, cerebralthrombosis, thromboembolic complications of surgery (such as joint orhip replacement), and peripheral arterial occlusion.

Any disease or condition that is associated with increased PAI-1activity or expression in a subject can be treated using substitutedheteroaryl benzofuran acids. Exemplary diseases and conditions includestroke, e.g., stroke associated with or resulting from atrialfibrillation; diseases associated with extracellular matrix accumulationincluding, but not limited to, renal fibrosis, chronic obstructivepulmonary disease, polycystic ovary syndrome, restenosis, renovasculardisease, and organ transplant rejection; diseases associated withneoangiogenesis, including, but not limited to, diabetic retinopathy;Alzheimer's disease, e.g., by increasing or normalizing levels ofplasmin concentration in a subject; and myelofibrosis with myeloidmetaplasia, e.g., by regulating stromal cell hyperplasia and increasesin extracellular matrix proteins.

The compounds of the present invention can be used to treat, forexample, diabetic nephropathy and renal dialysis associated withnephropathy; malignancies or cancers, including, but not limited to,leukemia, breast cancer and ovarian cancer; tumors, including, but notlimited to, liposarcomas and epithelial tumors; septicemia; obesity;insulin resistance; proliferative diseases, including, but not limitedto, psoriasis; conditions associated with abnormal coagulationhomeostasis; low grade vascular inflammation; cerebrovascular diseases;hypertension; dementia; osteoporosis; arthritis; respiratory diseases,such as asthma; heart failure; arrhythmia; angina, including, but notlimited to, angina pectoris; atherosclerosis and sequelae; kidneyfailure; multiple sclerosis; osteoporosis; osteopenia; dementia;peripheral vascular disease; peripheral arterial disease; acute vascularsyndromes; microvascular diseases including, but not limited to,nephropathy, neuropathy, retinopathy and nephrotic syndrome;hypertension; Type I and II diabetes and related diseases;hyperglycemia; hyperinsulinemia; malignant lesions; premalignantlesions; gastrointestinal malignancies; coronary heart disease,including, but not limited to, primary and secondary prevention ofmyocardial infarction, stable and unstable angina, primary prevention ofcoronary events, and secondary prevention of cardiovascular events; andinflammatory diseases, including, but not limited to, septic shock andthe vascular damage associated with infections.

The compounds of the present invention can also be administered to asubject in combination with a second therapeutic agent, including, butnot limited to, prothrombolytic, fibrinolytic, and anticoagulant agents,or in conjunction with other therapies, for example, proteaseinhibitor-containing highly active antiretroviral therapy (HAART) forthe treatment of diseases which originate from fibrinolytic impairmentand hyper-coagulability of HIV-1 infected patients. In certainembodiments, the compounds of the present invention can be administeredin conjunction with and/or following processes or procedures involvingmaintaining blood vessel patency, including, but not limited to,vascular surgery, vascular graft and stent patency, organ, tissue andcell implantation and transplantation. The compounds of the presentinvention can also be used for the treatment of blood and blood productsused in dialysis, blood storage in the fluid phase, especially ex vivoplatelet aggregation. The compounds of the present invention can also beadministered to a subject as a hormone replacement agent or to reduceinflammatory markers or C-reactive protein. The compounds can beadministered to improve coagulation homeostasis, to improve endothelialfunction, or as a topical application for wound healing, e.g., theprevention of scarring. The compounds of the present invention can beadministered to a subject in order to reduce the risk of undergoing amyocardial revascularization procedure. The present compounds can alsobe added to human plasma during the analysis of blood chemistry inhospital settings to determine the fibrinolytic capacity thereof. Incertain embodiments, the compounds of the present invention can be usedas imaging agents for the identification of metastatic cancers.

C. Synthesis of Substituted Heteroaryl Benzofuran Acids

Compounds of the present invention can be prepared by those skilled inthe art of organic synthesis employing conventional methods that utilizereadily available reagents and starting materials. Representativecompounds of the present invention can be prepared using the followingsynthetic schemes. The skilled practitioner will know how to make use ofvariants of these process steps, which in themselves are well known inthe art.

In certain embodiments of the present invention, substituted heteroarylbenzofuran acids can be prepared using scheme 1:

wherein R₂, R₃, and R₄ are defined as above for Formula 1 or 2 and R₈ ishydrogen, alkyl, cycloalkyl, perfluoroalkyl, alkenyl, alkynyl,arylalkenyl, arylalkynyl, or aryl.

In Scheme I, reaction of halo-hydroxy-quinolines 2 with methyl iodide inDMF under basic conditions afforded the methyl ether 2a. Coupling of 2awith benzofuran boronic acids in presence of Pd(OAc)₂, KF, and2-(di-t-butyl phosphino)-biphenyl in a solvent such as THF (J. P. Wolfe;S. L. Buchwald. Angew. Chem. Int. Ed., 38, 1999, 2413-2416) affordedcompounds 3. Bromination of the 3-position of the benzofuran usingbromine in acetic acid in the presence of sodium acetate afforded bromobenzofuran derivatives 4. Coupling of 4 with 1-alkenes under standardHeck reaction condition followed by hydrogenation afforded the alkylbenzofuran derivatives 5. Conversion of compounds 5 to the desiredderivatives 1a was carried out in three steps. Alkylation withbromoacetonitrile using a base such as potassium carbonate or cesiumcarbonate in a solvent such as acetone to give the correspondingnitriles. Conversion of the nitriles to the tetrazole derivatives 1a wascarried out by reacting with sodium azide in the presence of ammoniumchloride in a solvent such as DMF at a temperature of 80-100° C.Alternatively, bromo benzofuran derivatives 4 were reacted with arylboronic acids to yield the aryl benzofuran derivatives 6 which wereconverted to the desired derivatives 1b.

D. Substituted Heteroaryl Benzofuran Acids as PharmaceuticalCompositions

The present invention provides substituted heteroaryl benzofuran acidsas pharmaceuticals. In a preferred embodiment, the substitutedheteroaryl benzofuran acids are formulated as pharmaceuticals to treatdiseases associated with increased PAI-1 activity, e.g., by inhibitingPAI-1 activity in a subject.

In general, substituted heteroaryl benzofuran acids can be administeredas pharmaceutical compositions by any method known in the art foradministering therapeutic drugs including oral, buccal, topical,systemic (e.g., transdermal, intranasal, or by suppository), orparenteral (e.g., intramuscular, subcutaneous, or intravenousinjection). Compositions can take the form of tablets, pills, capsules,semisolids, powders, sustained release formulations, solutions,suspensions, emulsions, syrups, elixirs, aerosols, or any otherappropriate compositions; and comprise at least one compound of thisinvention in combination with at least one pharmaceutically acceptableexcipient. Suitable excipients are well known to persons of ordinaryskill in the art, and they, and the methods of formulating thecompositions, can be found in such standard references as Alfonso A R:Remington's Pharmaceutical Sciences, 17th ed., Mack Publishing Company,Easton Pa., 1985. Suitable liquid carriers, especially for injectablesolutions, include water, aqueous saline solution, aqueous dextrosesolution, and glycols. In some embodiments of the present invention,substituted heteroaryl benzofuran acids suitable for use in the practiceof this invention will be administered either singly or in combinationwith at least one other compound of this invention. Substitutedheteroaryl benzofuran acids suitable for use in the practice of thepresent invention can also be administered with at least one otherconventional therapeutic agent for the disease being treated.

Aqueous suspensions of the invention can contain an heteroarylbenzofuran acid in admixture with excipients suitable for themanufacture of aqueous suspensions. Such excipients can include asuspending agent, such as sodium carboxymethylcellulose,methylcellulose, hydroxypropylmethylcellulose, sodium alginate,polyvinylpyrrolidone, gum tragacanth and gum acacia, and dispersing orwetting agents such as a naturally occurring phosphatide (e.g.,lecithin), a condensation product of an alkylene oxide with a fatty acid(e.g., polyoxyethylene stearate), a condensation product of ethyleneoxide with a long chain aliphatic alcohol (e.g., heptadecaethyleneoxycetanol), a condensation product of ethylene oxide with a partialester derived from a fatty acid and a hexitol (e.g., polyoxyethylenesorbitol mono-oleate), or a condensation product of ethylene oxide witha partial ester derived from fatty acid and a hexitol anhydride (e.g.,polyoxyethylene sorbitan mono-oleate). The aqueous suspension can alsocontain one or more preservatives such as ethyl or n-propylp-hydroxybenzoate, one or more coloring agents, one or more flavoringagents, and one or more sweetening agents, such as sucrose, aspartame orsaccharin. Formulations can be adjusted for osmolarity.

Oil suspensions can be formulated by suspending a heteroaryl benzofuranacid in a vegetable oil, such as arachis oil, olive oil, sesame oil orcoconut oil, or in a mineral oil such as liquid paraffin; or a mixtureof these. The oil suspensions can contain a thickening agent, such asbeeswax, hard paraffin or cetyl alcohol. Sweetening agents can be addedto provide a palatable oral preparation, such as glycerol, sorbitol orsucrose. These formulations can be preserved by the addition of anantioxidant such as ascorbic acid. As an example of an injectable oilvehicle, see Minto, J. Pharmacol. Exp. Ther. 281:93-102, 1997. Thepharmaceutical formulations of the invention can also be in the form ofoil-in-water emulsions. The oily phase can be a vegetable oil or amineral oil, described above, or a mixture of these. Suitableemulsifying agents include naturally-occurring gums, such as gum acaciaand gum tragacanth, naturally occurring phosphatides, such as soybeanlecithin, esters or partial esters derived from fatty acids and hexitolanhydrides, such as sorbitan mono-oleate, and condensation products ofthese partial esters with ethylene oxide, such as polyoxyethylenesorbitan mono-oleate. The emulsion can also contain sweetening agentsand flavoring agents, as in the formulation of syrups and elixirs. Suchformulations can also contain a demulcent, a preservative, or a coloringagent.

The compound of choice, alone or in combination with other suitablecomponents, can be made into aerosol formulations (i.e., they can be“nebulized”) to be administered via inhalation. Aerosol formulations canbe placed into pressurized acceptable propellants, such asdichlorodifluoromethane, propane, nitrogen, and the like.

Formulations suitable for parenteral administration, such as, forexample, by intraarticular (in the joints), intravenous, intramuscular,intradermal, intraperitoneal, and subcutaneous routes, include aqueousand non-aqueous, isotonic sterile injection solutions, which can containantioxidants, buffers, bacteriostats, and solutes that render theformulation isotonic with the blood of the intended recipient, andaqueous and non-aqueous sterile suspensions that can include suspendingagents, solubilizers, thickening agents, stabilizers, and preservatives.Among the acceptable vehicles and solvents that can be employed arewater and Ringer's solution, an isotonic sodium chloride. In addition,sterile fixed oils can conventionally be employed as a solvent orsuspending medium. For this purpose any bland fixed oil can be employedincluding synthetic mono- or diglycerides. In addition, fatty acids suchas oleic acid can likewise be used in the preparation of injectables.These solutions are sterile and generally free of undesirable matter.Where the compounds are sufficiently soluble they can be dissolveddirectly in normal saline with or without the use of suitable organicsolvents, such as propylene glycol or polyethylene glycol. Dispersionsof the finely divided compounds can be made-up in aqueous starch orsodium carboxymethyl cellulose solution, or in suitable oil, such asarachis oil. These formulations can be sterilized by conventional, wellknown sterilization techniques. The formulations can containpharmaceutically acceptable auxiliary substances as required toapproximate physiological conditions such as pH adjusting and bufferingagents, toxicity adjusting agents, e.g., sodium acetate, sodiumchloride, potassium chloride, calcium chloride, sodium lactate and thelike. The concentration of substituted heteroaryl benzofuran acids inthese formulations can vary widely, and will be selected primarily basedon fluid volumes, viscosities, body weight, and the like, in accordancewith the particular mode of administration selected and the patient'sneeds. For IV administration, the formulation can be a sterileinjectable preparation, such as a sterile injectable aqueous oroleaginous suspension. This suspension can be formulated according tothe known art using those suitable dispersing or wetting agents andsuspending agents. The sterile injectable preparation can also be asterile injectable solution or suspension in a nontoxicparenterally-acceptable diluent or solvent, such as a solution of1,3-butanediol. The formulations of commends can be presented inunit-dose or multi-dose sealed containers, such as ampules and vials.

Injection solutions and suspensions can be prepared from sterilepowders, granules, and tablets of the kind previously described.

Substituted heteroaryl benzofuran acids suitable for use in the practiceof this invention can be administered orally. The amount of a compoundof the present invention in the composition can vary widely depending onthe type of composition, size of a unit dosage, kind of excipients, andother factors well known to those of ordinary skill in the art. Ingeneral, the final composition can comprise, for example, from 0.000001percent by weight (% w) to 10% w of the substituted heteroarylbenzofuran acid, preferably 0.00001% w to 1% w, with the remainder beingthe excipient or excipients.

Pharmaceutical formulations for oral administration can be formulatedusing pharmaceutically acceptable carriers well known in the art indosages suitable for oral administration. Such carriers enable thepharmaceutical formulations to be formulated in unit dosage forms astablets, pills, powder, dragees, capsules, liquids, lozenges, gels,syrups, slurries, suspensions, etc. suitable for ingestion by thepatient. Formulations suitable for oral administration can consist of(a) liquid solutions, such as an effective amount of the packagednucleic acid suspended in diluents, such as water, saline or PEG 400;(b) capsules, sachets or tablets, each containing a predetermined amountof the active ingredient, as liquids, solids, granules or gelatin; (c)suspensions in an appropriate liquid; and (d) suitable emulsions.

Pharmaceutical preparations for oral use can be obtained throughcombination of the compounds of the present invention with a solidexcipient, optionally grinding a resulting mixture, and processing themixture of granules, after adding suitable additional compounds, ifdesired, to obtain tablets or dragee cores. Suitable solid excipientsare carbohydrate or protein fillers and include, but are not limited tosugars, including lactose, sucrose, mannitol, or sorbitol; starch fromcorn, wheat, rice, potato, or other plants; cellulose such as methylcellulose, hydroxymethyl cellulose, hydroxypropylmethyl-cellulose orsodium carboxymethylcellulose; and gums including arabic and tragacanth;as well as proteins such as gelatin and collagen. If desired,disintegrating or solubilizing agents can be added, such as thecross-linked polyvinyl pyrrolidone, agar, alginic acid, or a saltthereof, such as sodium alginate. Tablet forms can include one or moreof lactose, sucrose, mannitol, sorbitol, calcium phosphates, cornstarch, potato starch, microcrystalline cellulose, gelatin, colloidalsilicon dioxide, talc, magnesium stearate, stearic acid, and otherexcipients, colorants, fillers, binders, diluents, buffering agents,moistening agents, preservatives, flavoring agents, dyes, disintegratingagents, and pharmaceutically compatible carriers. Lozenge forms cancomprise the active ingredient in a flavor, e.g., sucrose, as well aspastilles comprising the active ingredient in an inert base, such asgelatin and glycerin or sucrose and acacia emulsions, gels, and the likecontaining, in addition to the active ingredient, carriers known in theart.

The substituted heteroaryl benzofuran acids of the present invention canalso be administered in the form of suppositories for rectaladministration of the drug. These formulations can be prepared by mixingthe drug with a suitable non-irritating excipient which is solid atordinary temperatures but liquid at the rectal temperatures and willtherefore melt in the rectum to release the drug. Such materials arecocoa butter and polyethylene glycols.

The compounds of the present invention can also be administered byintranasal, intraocular, intravaginal, and intrarectal routes includingsuppositories, insufflation, powders and aerosol formulations (forexamples of steroid inhalants, see Rohatagi, J. Clin. Pharmacol.35:1187-1193, 1995; Tjwa, Ann. Allergy Asthma Immunol. 75:107-111,1995).

The substituted heteroaryl benzofuran acids of the present invention canbe delivered transdermally, by a topical route, formulated as applicatorsticks, solutions, suspensions, emulsions, gels, creams, ointments,pastes, jellies, paints, powders, and aerosols.

Encapsulating materials can also be employed with the compounds of thepresent invention and the term “composition” is intended to include theactive ingredient in combination with an encapsulating material as aformulation, with or without other carriers. For example, the compoundsof the present invention can also be delivered as microspheres for slowrelease in the body. In one embodiment, microspheres can be administeredvia intradermal injection of drug-containing microspheres, which slowlyrelease subcutaneously (see Rao, J. Biomater Sci. Polym. Ed. 7:623-645,1995; as biodegradable and injectable gel formulations (see, e.g., Gao,Pharm. Res. 12:857-863, 1995); or, as microspheres for oraladministration (see, e.g., Eyles, J. Pharm. Pharmacol. 49:669-674,1997). Both transdermal and intradermal routes afford constant deliveryfor weeks or months. Cachets can also be used in the delivery of thecompounds of the present invention, e.g., anti-atheroscleroticmedicaments.

In another embodiment, the compounds of the present invention can bedelivered by the use of liposomes which fuse with the cellular membraneor are endocytosed, i.e., by employing ligands attached to the liposome,or attached directly to the oligonucleotide, that bind to surfacemembrane protein receptors of the cell resulting in endocytosis. Byusing liposomes, particularly where the liposome surface carries ligandsspecific for target cells, or are otherwise preferentially directed to aspecific organ, one can focus the delivery of the compound into thetarget cells in vivo. (See, e.g., Al-Muhammed, J. Microencapsul.13:293-306, 1996; Chonn, Curr. Opin. Biotechnol. 6:698-708, 1995; Ostro,Am. J. Hosp. Pharm. 46:1576-1587, 1989).

In other cases, the preferred preparation can be a lyophilized powder in1 mM-50 mM histidine, 0.1%-2% sucrose, 2%-7% mannitol at a pH range of4.5 to 5.5, that is combined with buffer prior to use.

A pharmaceutical composition of the invention can optionally contain, inaddition to a substituted heteroaryl benzofuran acid, at least one othertherapeutic agent useful in the treatment of a disease or conditionassociated with increased PAI-1 activity.

The pharmaceutical compositions are generally formulated as sterile,substantially isotonic and in full compliance with all GoodManufacturing Practice (GMP) regulations of the U.S. Food and DrugAdministration.

E. Determining Dosage Regimens for Substituted Heteroaryl BenzofuranAcids

The present invention provides methods of inhibiting PAI-1 activity in amammal for the treatment of diseases and conditions associated withincreased PAI-1 activity using substituted heteroaryl benzofuran acids.

For treatment purposes, the compositions or compounds disclosed hereincan be administered to the subject in a single bolus delivery, viacontinuous delivery (e.g., continuous transdermal, mucosal, orintravenous delivery) over an extended time period, or in a repeatedadministration protocol (e.g., by an hourly, daily or weekly, repeatedadministration protocol). The pharmaceutical formulations of the presentinvention can be administered, for example, one or more times daily, 3times per week, or weekly. In an exemplary embodiment of the presentinvention, the pharmaceutical formulations of the present invention areorally administered once or twice daily.

In this context, a therapeutically effective dosage of the biologicallyactive agent(s) can include repeated doses within a prolonged treatmentregimen that will yield clinically significant results to alleviate oneor more symptoms or detectable conditions associated with increasedPAI-1 activity. Determination of effective dosages in this context istypically based on animal model studies followed up by human clinicaltrials and is guided by determining effective dosages and administrationprotocols that significantly reduce the occurrence or severity oftargeted exposure symptoms or conditions in the subject. Suitable modelsin this regard include, for example, murine, rat, porcine, feline,non-human primate, and other accepted animal model subjects known in theart. Alternatively, effective dosages can be determined using in vitromodels (e.g., immunologic and histopathologic assays). Using suchmodels, only ordinary calculations and adjustments are typicallyrequired to determine an appropriate concentration and dose toadminister a therapeutically effective amount of the biologically activeagent(s) (e.g., amounts that are intranasally effective, transdermallyeffective, intravenously effective, or intramuscularly effective toelicit a desired response). In alternative embodiments, an “effectiveamount” or “therapeutically effective dose” of the biologically activeagent(s) will simply inhibit or enhance one or more selected biologicalactivity(ies) correlated with a disease or condition, as set forthabove, for either therapeutic or diagnostic purposes.

The actual dosage of biologically active agents will of course varyaccording to factors such as the extent of exposure and particularstatus of the subject (e.g., the subject's age, size, fitness, extent ofsymptoms, susceptibility factors, etc), time and route ofadministration, as well as other drugs or treatments being administeredconcurrently. Dosage regimens can be adjusted to provide an optimumprophylactic or therapeutic response. By “therapeutically effectivedose” herein is meant a dose that produces effects for which it isadministered. More specifically, a therapeutically effective dose of thecompound(s) of the invention preferably alleviates symptoms,complications, or biochemical indicia of diseases associated withincreased PAI-1 activity. The exact dose will depend on the purpose ofthe treatment, and will be ascertainable by one skilled in the art usingknown techniques (see, e.g., Lieberman, Pharmaceutical Dosage Forms(Vols. 1-3, 1992); Lloyd, 1999, The Art, Science, and Technology ofPharmaceutical Compounding; and Pickar, 1999, Dosage Calculations). Atherapeutically effective dose is also one in which any toxic ordetrimental side effects of the active agent is outweighed in clinicalterms by therapeutically beneficial effects. It is to be further notedthat for each particular subject, specific dosage regimens should beevaluated and adjusted over time according to the individual need andprofessional judgment of the person administering or supervising theadministration of the compound.

In an exemplary embodiment of the present invention, unit dosage formsof the compounds are prepared for standard administration regimens. Inthis way, the composition can be subdivided readily into smaller dosesat the physicians direction. For example, unit dosages can be made up inpacketed powders, vials or ampoules and preferably in capsule or tabletform. The active compound present in these unit dosage forms of thecomposition can be present in an amount of, for example, from about onegram to about fifteen grams or more, for single or multiple dailyadministration, according to the particular need of the patient. Byinitiating the treatment regimen with a minimal daily dose of about onegram, the blood levels of PAI-1 and the patients symptomatic reliefanalysis can be used to determine whether a larger or smaller dose isindicated. Effective administration of the compounds of this inventioncan be given at an oral dose of, for example, from about 0.1 mg/kg/dayto about 1,000 mg/kg/day. Preferably, administration will be from about10/mg/kg/day to about 600 mg/kg/day, more preferably from about 25 toabout 200 mg/kg/day, and even more preferably from about 50 mg/kg/day toabout 100 mg/kg/day. In some embodiments, a daily dosage of from about 1mg/kg to about 250 mg/kg is provided.

In certain embodiments, the present invention is directed to prodrugs ofcompounds of formulas 1 or 2. The term “prodrug,” as used herein, meansa compound that is convertible in vivo by metabolic means (e.g. byhydrolysis) to a compound of formulas 1 or 2. Various forms of prodrugsare known in the art such as those discussed in, for example, Bundgaard,(ed.), Design of Prodrugs, Elsevier (1985); Widder, et al. (ed.),Methods in Enzymology, vol. 4, Academic Press (1985); Krogsgaard-Larsen,et al., (ed). “Design and Application of Prodrugs, Textbook of DrugDesign and Development, Chapter 5, 113-191 (1991), Bundgaard, et al.,Journal of Drug Delivery Reviews, 8:1-38 (1992), Bundgaard, J. ofPharmaceutical Sciences, 77:285 et seq. (1988); and Higuchi and Stella(eds.) Prodrugs as Novel Drug Delivery Systems, American ChemicalSociety (1975).

F. Kits

After a pharmaceutical comprising a substituted heteroaryl benzofuranacid has been formulated in a suitable carrier, it can be placed in anappropriate container and labeled for treatment of a PAI-1 relateddisorder, e.g., leukemia. Additionally, another pharmaceuticalcomprising at least one other therapeutic agent useful in the treatmentof the PAI-1 related disorder can be placed in the container as well andlabeled for treatment of the indicated disease. Alternatively, a singlepharmaceutical comprising a substituted heteroaryl benzofuran acid andat least one other therapeutic agent useful in the treatment of a PAI-1related disorder can be placed in an appropriate container and labeledfor treatment. For administration of pharmaceuticals comprisingsubstituted heteroaryl benzofuran acids and of pharmaceuticalscomprising, in a single pharmaceutical, substituted heteroarylbenzofuran acids and at least one other therapeutic agent useful in thetreatment of a PAI-1 related disorder, such labeling would include, forexample, instructions concerning the amount, frequency and method ofadministration. Similarly, for administration of multiplepharmaceuticals provided in the container, such labeling would include,for example, instructions concerning the amount, frequency and method ofadministration of each pharmaceutical.

EXAMPLES

The syntheses of compounds I-5 are described in examples 1-5respectively.

Example 1 Synthesis of6-(1-Benzofuran-2-yl)-2-(trifluoromethyl)quinolin-4-yl1H-tetraazol-5-ylmethyl ether

Step 1: Synthesis of6-(1-Benzofuran-2-yl)-4-hydroxy-2-(trifluoromethyl)quinoline: To a flaskwith 6-bromo-4-hydroxy-2-trifluoromethyl-quinoline (1.00 g, 3.42 mmol)was added K₂CO₃ (8.55 mL, 1.0 M in H₂O, 8.55 mmol) followed by dioxane(85.5 mL). Benzofuran-2-boronic acid (0.665 g, 4.10 mmol) was then addedfollowed by PdCl₂(dppf) (0.028 g, 0.0342 mmol). The reaction mixture wasstirred at room temperature for 0.5 hour and then heated to 65° C. for18 hours. After concentration, the reaction was diluted with EtOAc (200mL). The organic layer was washed with 1 N HCl, (20 mL), sat. aq. NaHCO₃(20 mL), and brine (20 mL) and then dried (MgSO₄). After concentration,the residue was recrystallized from EtOAc to afford the product (0.770g, 68%) as a solid. Mass spectrum (+ESI, [M+H]⁺) m/z 330. ¹H NMR (500MHz, DMSO-d₆): δ 8.68 (s, 1H), 8.37 (d, 1H, J=9.3 Hz), 8.07 (d, 1H,J=9.3 Hz), 7.65-7.75 (m, 3H), 7.38 (t, 1H, J=7.7 Hz), 7.30 (t, 1H, J=7.7Hz), and 7.06 ppm (s, 1H).

Step 2: Synthesis of{[6-(1-Benzofuran-2-yl)-2-(trifluoromethyl)quinolin-4-yl]oxy}acetonitrile:To a stirred solution of6-(1-benzofuran-2-yl)-4-hydroxy-2-(trifluoromethyl)quinoline (0.770 g,2.34 mmol) in acetone (40 mL) at room temperature was added Cs₂CO₃(0.839 g, 2.57 mmol) followed by bromoacetonitrile (0.326 mL, 4.68 mmol)dropwise. The reaction was stirred at this temperature for 18 hours andthen diluted with EtOAc (200 mL). The organic layer was washed with 1 NHCl (20 mL), sat. aq. NaHCO₃ (20 mL), and brine (20 mL) and then dried(MgSO₄). After concentration, the residue was recrystallized fromEtOAc:Hexane to afford the product (0.711 g, 82%) as a off-white solid,mp 174-176° C. Mass spectrum (+APCI, [M+H]⁺) m/z 369. ¹H NMR (500 MHz,DMSO-d₆): δ 8.61 (d, 1H, J=2.0), 8.48 (dd, 1H, J=2.0, 8.8 Hz), 8.23 (d,1H, J=8.8 Hz), 7.78 (s, 1H), 7.70-7.75 (m, 2H), 7.67 (s, 1H), 7.39 (t,1H, J=7.2 Hz), 7.30 (t, 1H, J=7.6 Hz), and 5.70 ppm (s, 2H), Elementalanalysis for C₂₀H₁₁F₃N₂O₂: Calculated: C, 65.22; H, 3.01; N, 7.61.Found: C, 65.02; H, 2.87; N, 7.51.

Step 3: Synthesis of6-(1-Benzofuran-2-yl)-2-(trifluoromethyl)quinolin-4-yl1H-tetraazol-5-ylmethyl ether: To a stirred solution of{[6-(1-benzofuran-2-yl)-2-(trifluoromethyl)quinolin-4-yl]oxy}acetonitrile(0.200 g, 0.543 mmol) in DMF (10 mL) at room temperature was addedsodium azide (0.177 g, 2.72 mmol) followed by NH₄Cl (0.145 g, 2.72mmol). The reaction was heated to 100° C. for 2 hours. The mixture wasconcentrated and diluted with 2 N HCl (˜5 mL). This mixture was stirredat room temperature for 1 hour. The solid which formed was filtered andthen washed with excess H₂O and hexane and dried under high vacuum toafford the product (0.173 g, 78%) as a yellow solid, mp >225° C. (dec.),mp >225° C. (dec.). Mass spectrum (+APCI, [M+H]⁺) m/z 412. ¹H NMR (500MHz, DMSO-d₆): δ 15.44-17.96 (br s, 1H), 8.72 (d, 1H, J=2.0 Hz), 8.46(dd, 1H, J=2.0, 9.0 Hz), 8.21 (d, 1H, J=8.8 Hz), 7.70-7.75 (m, 3H), 7.76(s, 1H), 7.38 (t, 1H, J=7.7 Hz), 7.30 (t, 1H, J=7.9 Hz), and 6.04 ppm(s, 2H). Elemental analysis for C₂₀H₁₂F₃N₅O₂.0.5H₂O:Calculated: C,57.15; H, 3.12; N, 16.66. Found: C, 57.43; H, 2.72; N, 16.53.

Example 2 Synthesis of6-(3-Pentyl-1-benzofuran-2-yl)-4-(1H-tetraazol-5-ylmethoxy)-2-(trifluoromethyl)-quinoline

Step 1: Synthesis of6-(1-Benzofuran-2-yl)-4-methoxy-2-(trifluoromethyl)quinoline: To astirring solution of6-(1-benzofuran-2-yl)-4-hydroxy-2-(trifluoromethyl)-quinoline (1.33 g,4.04 mmol) in DMF (40 mL) at room temperature was added K₂CO₃ (0.670 g,4.85 mmol) followed by methyl iodide (0.276 mL, 4.44 mmol) drop-wise.The reaction mixture was stirred for 4 hours then concentrated and theresidue was diluted with EtOAc (200 mL). The organic layer was washedwith H₂O (20 mL), sat. aq. NaHCO₃ (20 mL), and brine (20 mL) and thendried (Na₂SO₄). After concentration, the residue was purified on aBiotage Flash 40 apparatus (5-15% EtOAc:petroleum ether) to afford theproduct (0.930 g) as a solid. Mass spectrum (+ESI, [M+H]⁺) m/z 344. ¹HNMR (500 MHz, DMSO-d₆): δ 8.67 (s, 1H), 8.46 (d, 1H, J=9.1 Hz), 8.19 (d,1H, J=9.1 Hz), 7.68-7.79 (m, 3H), 7.48 (s, 1H), 7.40 (t, 1H, J=7.8 Hz),7.32 (t, 1H, J=7.8 Hz), and 4.26 ppm (s, 3H).

Step 2: Synthesis of6-(3-Bromo-1-benzofuran-2-yl)-4-methoxy-2-(trifluoromethyl)quinoline: Toa stirred solution of6-(1-benzofuran-2-yl)-4-methoxy-2-(trifluoromethyl)quinoline (0.930 g,2.71 mmol) in HOAc (30 mL) at 0° C. was added KOAc (0.319 g, 3.25 mmol).The reaction was stirred at this temperature for 10 minutes, and then asolution of Br₂ (0.154 mL, 2.98 mmol) in HOAc (5 mL) was added dropwiseto it over a period of 10 minutes. The reaction mixture was allowed towarm to room temperature and stirred for 2.5 hours. The reaction mixturewas then diluted with H₂O (10 mL). The solid that formed was filteredoff and washed with excess H₂O and hexane. This solid was thenrecrystallized from EtOAc:hexane to afford the product (0.750 g, 66%) asa solid. Mass spectrum (+ESI, [M+H]⁺) m/z 422/424. ¹H NMR (500 MHz,DMSO-d₆): δ 8.95 (s, 1H), 8.65 (d, 1H, J=8.9 Hz), 8.29 (d, 1H, J=8.9Hz), 7.80 (d, 1H, J=8.1 Hz), 7.66 (d, 1H, J=8.1 Hz), 7.36-7.58 (m, 3H),and 4.26 ppm (s, 3H).

Step 3: Synthesis of6-(3-Pentyl-1-benzofuran-2-yl)-4-methoxy-2-(trifluoromethyl)quinoline:The mixture of6-(3-bromo-1-benzofuran-2-yl)-4-methoxy-2-(trifluoromethyl)quinoline(0.747 g, 1.77 mmol) 1-pentene (0.233 mL, 2.12 mmol), Et₃N (7.4 mL, 53.1mmol), Pd(OAc)₂ (0.004 g, 0.0177 mmol) and P(o-tol)₃ (0.022 g, 0.0708mmol) in DMF (7.4 mL) was heated in sealed tube at 100° C. for 18 hours.The mixture was then cooled to room temperature and quenched with H₂O(20 mL) and diluted with EtOAc (200 mL). The organic layer was washedwith H₂O (20 mL) and brine (20 mL) and then dried (Na₂SO₄). Afterconcentration, the residue was purified by on a Biotage Flash 40apparatus (5-10% EtOAc:petroleum ether) to afford6-(3-pent-1-enyl-benzofuran-2-yl)-4-methoxy-2-(trifluoromethyl)quinoline(0.273 g) as an oil. Mass spectrum (+ESI, [M+H]⁺) m/z 412. To a solutionof this intermediate in EtOAc:MeOH (1:1, 16 mL) at room temperature wasadded Pd/C (0.041 g, 1.91 mmol). This mixture was stirred under anatmosphere of H₂ for 5 hours. The mixture was then filtered throughcelite, and the bed of celite was washed with excess EtOAc:MeOH (1:1).After concentration, the residue was dried under high vacuum to affordthe product (0.258 g) as an oil. Mass spectrum (+ESI, [M+H]⁺) m/z 414.¹H NMR (500 MHz, DMSO-d₆): δ 8.58 (s, 1H), 8.18-8.35 (m, 2H), 7.74 (d,1H, J=8.7 Hz), 7.68 (d, 1H, J=8.7 Hz), 7.50 (s, 1H), 7.27-7.46 (m, 2H),4.26 (s, 3H), 3.05 (t, 2H, J=8.3 Hz), 1.71-1.87 (m, 2H), 1.30-1.53 (m,4H), and 0.89 ppm (t, 3H, J=7.5 Hz).

Step 4: Synthesis of{[6-(3-Pentyl-1-benzofuran-2-yl)-2-(trifluoromethyl)quinolin-4-yl]oxy}acetonitrile:To a stirred solution of6-(3-pentyl-1-benzofuran-2-yl)-4-methoxy-2-(trifluoromethyl) quinoline(0.258 g, 0.624 mmol) in CH₂Cl₂ (10 mL) cooled to −78° C. was added BBr₃(1.68 mL, 1.0 M in CH₂Cl₂, 1.68 mmol) dropwise. The reaction was stirredat this temperature for 0.5 hour and then warmed to room temperature for18 hours. The reaction mixture was then heated at reflux for 4 hours. Atthis point, BCl₃ (1.68 mL, 1.0 M in CH₂Cl₂, 1.68 mmol) and Bu₄N⁺I⁻ (1.24g, 3.37 mmol) were added and the mixture was allowed to stir at roomtemperature for 3 days. The reaction mixture was then quenched with MeOH(˜5 mL) followed by dilution with H₂O (20 mL) and EtOAc (200 mL). Theorganic layer was washed with brine (20 mL) and then dried (Na₂SO₄).After concentration, the residue was dried under high vacuum to affordthe product (0.247 g, 99%) as an oil. Mass spectrum (+ESI, (M+H]⁺) m/z400. Reaction of this intermediate[6-(3-Pentyl-1-benzofuran-2-yl)-4-hydroxy-2-(trifluoromethyl)quinoline]with bromoacetonitrile according to the procedure described in Step 2 ofExample 1; afforded the title compound as a solid. Mass spectrum (+ESI,[M+H]⁺) m/z 439. ¹H NMR (500 MHz, DMSO-d₆): δ 8.60 (s, 1H), 8.18-8.33(m, 2H), 7.55-7.77 (m, 3H), 7.29-7.42 (m, 2H), 5.68 (s, 2H), 3.04 (t,2H, J=8.2 Hz), 1.73-1.86 (m, 2H), 1.31-1.51 (m, 4H), and 0.89 ppm (t,3H, J=7.6 Hz).

Step 5: Synthesis of6-(3-Pentyl-1-benzofuran-2-yl)-4-(1H-tetraazol-5-ylmethoxy)-2-(trifluoromethyl)-quinoline:The title compound was prepared as a white solid (0.054 g, 99%) from{[6-(3-pentyl-1-benzofuran-2-yl)-2-(trifluoromethyl)quinolin-4-yl]oxy}acetonitrilefollowing the procedure described in Step 3 of Example 1, mp 207-209° C.(dec.). Mass spectrum (+ESI, [M+H]⁺) m/z 482. ¹H NMR (500 MHz, DMSO-d₆):δ 16.30-17.90 (br s, 1H), 8.60 (d, 1H, J=1.7 Hz), 8.32 (dd, 1H, J=2.0,8.9 Hz), 8.27 (d, 1H, J=8.9 Hz), 7.72-7.76 (m, 2H), 7.66 (d, 1H, J=8.1Hz), 7.39 (t, 1H, J=7.6 Hz), 7.32 (t, 1H, J=7.8 Hz), 6.01 (s, 2H), 2.99(t, 2H, J=7.6 Hz), 1.64-1.72 (m, 2H), 1.19-1.33 (m, 4H), and 0.77 ppm(t, 3H, J=7.2 Hz), Elemental Analysis for C₂₅H₂₂F₃N₅O₂.0.25H₂O:Calculated: C, 61.79; H, 4.67; N, 14.41. Found: C, 61.91; H, 4.31; N,14.51.

Example 3 Synthesis of5-(1-Benzofuran-2-yl)-8-(1H-tetraazol-5-ylmethoxy)quinoline

Step 1: Synthesis of 5-Chloro-8-methoxy-quinoline: The title compoundwas prepared as a solid (0.768 g, 71%) from 5-chloro-8-hydroxy-quinolinefollowing the procedure described in Step 1 of Example 2. Mass spectrum(+ESI, [M+H]⁺) m/z 194. ¹H NMR (500 MHz, DMSO-d₆): δ 8.95 (d, 1H, J=2.1Hz), 8.51 (d, 1H, J=8.5 Hz), 7.68-7.76 (m, 2H), 7.21 (d, 1H, J=7.8 Hz),and 3.98 ppm (s, 3H).

Step 2: Synthesis of 5-(1-Benzofuran-2-yl)-8-methoxy-quinoline. To astirred solution of 5-chloro-8-methoxy-quinoline (0.330 g, 1.70 mmol) inTHF (2 mL) at room temperature was added benzofuran-2-boronic acid(0.413 g, 2.55 mmol) followed by KF (0.296 mL, 5.10 mmol), Pd(OAc)₂(0.0382 g, 0.170 mmol), and 2-(di-tert-butyl phosphino)biphenyl (0.101g, 0.340 mmol). The reaction mixture was stirred at room temperature for18 hours and then diluted with EtOAc (200 mL). The organic layer waswashed with H₂O (20 mL) and brine (20 mL) then dried (Na₂SO₄). Afterconcentration, the residue was purified on a Biotage Flash 40 apparatus(30-70% EtOAc:petroleum ether) to afford the product (0.291 g, 62%) as asolid. Mass spectrum (+ESI, [M+H]⁺) m/z 276. ¹H NMR (500 MHz, DMSO-d₆):δ 8.94 (d, 1H, J=2.2 Hz), 8.83 (d, 1H, J=9.4 Hz), 7.99 (d, 1H, J=8.6Hz), 7.66-7.77 (m, 3H), 7.27-7.41 (m, 4H), and 4.6 ppm (s, 3H).

Step 3: Synthesis of{[5-(1-Benzofuran-2-yl)-quinolin-8-yl]oxy}acetonitrile:5-(1-Benzofuran-2-yl)-8-hydroxy-quinoline was prepared as a solid (0.264g, 99%) from 5-(1-benzofuran-2-yl)-8-methoxy-quinoline following theprocedure described in the first part of Step 4 of Example 2. Massspectrum (+ESI, [M+H]⁺) m/z 262. Reaction of5-(1-benzofuran-2-yl)-8-hydroxy-quinoline with bromoacetonitrileaccording to the procedure described in Step 2 of Example 1 afforded thetitle compound as a solid (0.104 g, 34%). Mass spectrum (+ESI, [M+H]⁺)m/z 301. ¹H NMR (500 MHz, DMSO-d₆): δ 9.02 (d, 1H, J=1.4 Hz), 8.99 (d,1H, J=9.3 Hz), 8.07 (d, 1H, J=8.3 Hz), 7.68-7.81 (m, 3H), 7.54 (d, 1H,J=8.3 Hz), 7.28-7.46 (m, 3H), and 5.51 ppm (s, 2H).

Step 4: Synthesis of5-(1-Benzofuran-2-yl)-8-(1H-tetraazol-5-ylmethoxy)quinoline: The titlecompound was prepared as a orange solid (0.087 g, 73%) from{[5-(1-benzofuran-2-yl)-quinolin-8-yl]oxy}acetonitrile according to theprocedure described in Step 3 of Example 1, mp >240° C. (dec.). Massspectrum (+APCI, [M+H]⁺) m/z 344. ¹H NMR (500 MHz, DMSO-d₆): δ15.30-17.90 (br s, 1H), 8.99-9.05 (m, 2H), 8.07 (d, 1H, J=8.3 Hz), 7.81(dd, 1H, J=4.4, 8.5 Hz), 7.72-7.75 (m, 1H), 7.69 (d, 1H, J=7.8 Hz), 7.63(d, 1H, J=8.3 Hz), 7.40 (s, 1H), 7.38 (td, 1H, J=1.7, 8.3 Hz), 7.32 (td,1H, J=1.0, 7.3 Hz), and 5.83 ppm (s, 2H). Elemental Analysis forC₁₉H₁₃N₅O₂.HCl, Calculated: C, 60.09; H, 3.72; N, 18.44. Found: C,60.31; H, 3.59; N, 18.65.

Example 4 Synthesis of 5-(3-Phenyl-1-benzofuran-2-yl)-8-quinolinyl1H-tetraazol-5-ylmethyl ether

Step 1: Synthesis of 5-(3-Bromo-1-benzofuran-2-yl)-8-methoxy-quinoline:The title compound was prepared as a solid (0.963 g, 0.74%) from5-(1-benzofuran-2-yl)-8-methoxy-quinoline following the proceduredescribed in Step 2 of Example 2. Mass spectrum (+ESI, [M+H]⁺) m/z354/356. ¹H NMR (400 MHz, DMSO-d₆): δ 8.95 (d, 1H, J=2.2 Hz), 8.29 (d,1H, J=8.7 Hz), 7.92 (d, 1H, J=8.1 Hz), 7.76 (d, 1H, J=8.1 Hz), 7.58-7.72(m, 2H), 7.31-7.58 (m, 3H), and 4.07 ppm (s, 3H).

Step 2: Synthesis of 5-(3-Phenyl-1-benzofuran-2-yl)-8-methoxy-quinoline:The title compound was prepared as a solid (0.202 g, 68%) by thecoupling of 5-(3-bromo-1-benzofuran-2-yl)-8-methoxy-quinoline and phenylboronic acid according to the procedure described in Step 1 ofExample 1. Mass spectrum (+ESI, [M+H]⁺) m/z 352. ¹H NMR (400 MHz,DMSO-d₆): δ 8.87 (d, 1H, J=1.6 Hz), 8.13 (d, 1H, J=8.2 Hz), 7.73 (t, 2H,J=7.8 Hz), 7.68 (d, 1H, J=8.2 Hz), 7.38-7.52 (m, 3H), 7.21-7.38 (m, 6H),and 4.00 ppm (s, 3H).

Step 3: Synthesis of{[5-(3-Phenyl-1-benzofuran-2-yl)-quinolin-8-yl]oxy}acetonitrile:5-(3-Phenyl-1-benzofuran-2-yl)-8-hydroxy-quinoline was prepared as asolid (0.185 g, 99%) from5-(3-phenyl-1-benzofuran-2-yl)-8-methoxy-quinoline according to theprocedure described in the first part of Step 4 of Example 2; Massspectrum (+ESI, [M+H]⁺) m/z 338. Reaction of5-(3-phenyl-1-benzofuran-2-yl)-8-hydroxy-quinoline withbromoacetonitrile according to the procedure described in Step 2 ofExample 1 afforded the title compound as a solid (0.148 g, 74%). Massspectrum (+ESI, [M+H]⁺) m/z 377. ¹H NMR (400 MHz, DMSO-d₆): δ 8.90-8.96(m, 1H), 8.19 (d, 1H, J=8.1 Hz), 7.71-7.81 (m, 3H), 7.39-7.65 (m, 4H),7.25-7.39 (m, 5H), and 5.48 ppm (s, 2H).

Step 4: Synthesis of 5-(3-Phenyl-1-benzofuran-2-yl)-8-quinolinyl1H-tetraazol-5-ylmethyl ether: The title compound was prepared as ayellow solid (0.125 g, 84%) from{[5-(3-phenyl-1-benzofuran-2-yl)-quinolin-8-yl]oxy}acetonitrileaccording to the procedure described in Step 3 of Example 1, mp >273° C.(dec.). Mass spectrum (+ESI, [M+H]⁺) m/z 420. ¹H NMR (400 MHz, DMSO-d₆):δ 15.30-17.90 (br s, 1H), 8.96 (d, 1H, J=3.8 Hz), 8.38 (d, 1H, J=8.6Hz), 7.73-7.82 (m, 3H), 7.62 (dd, 1H, J=4.3, 8.6 Hz), 7.57 (d, 1H, J=8.2Hz), 7.47 (t, 1H, J=7.8 Hz), 7.40 (t, 1H, J=7.7 Hz), 7.31-7.37 (m, 4H),7.25-7.31 (m, 1H), and 5.80 ppm (s, 2H). Elemental Analysis forC₂₅H₁₇N₅O₂.HCl: Calculated: C, 65.86; H, 3.98; N, 15.36. Found: C,66.07; H, 4.02; N, 15.31.

Example 5 Synthesis of5-(3-Pentyl-1-benzofuran-2-yl)-8-(1H-tetraazol-5-ylmethoxy)quinoline

Step 1: Synthesis of 5-(3-Pentyl-1-benzofuran-2-yl)-8-methoxy-quinoline:The title compound was prepared as a solid (0.132 g, 41%) from5-(3-bromo-1-benzofuran-2-yl)-8-methoxy-quinoline according to theprocedure described in Step 3 of Example 2. Mass spectrum (+ESI, [M+H)⁺)m/z 346. ¹H NMR (500 MHz, DMSO-d₆): δ 8.92 (d, 1H, J=1.9 Hz), 8.13 (d,1H, J=8.8 Hz), 7.54-7.67 (m, 2H), 7.69-7.78 (m, 2H), 4.06 (s, 3H),7.28-7.42 (m, 3H), 2.67 (t, 2H, J=8.4 Hz), 1.02-1.20 (m, 4H), 1.51-1.66(m, 2H), and 0.67 ppm (t, 3H, J=6.9 Hz).

Step 2: Synthesis of{[5-(3-Pentyl-1-benzofuran-2-yl)-quinolin-8-yl]oxy}acetonitrile:5-(3-Pentyl-1-benzofuran-2-yl)-8-hydroxy-quinoline was prepared as asolid (0.113 g, 99%) from5-(3-pentyl-1-benzofuran-2-yl)-8-methoxy-quinoline according to theprocedure described in the first part of Step 4 of Example 2. Massspectrum (+ESI, [M+H)⁺) m/z 332. Reaction of5-(3-Pentyl-1-benzofuran-2-yl)-8-hydroxy-quinoline withbromoacetonitrile according to the procedure described in Step 2 ofExample 1 afforded the title compound as a solid (0.059 g, 48%). Massspectrum (+ESI, [M+H)⁺) m/z 371. ¹H NMR (500 MHz, DMSO-d₆): δ 8.94-9.02(m, 1H), 8.20 (d, 1H, J=9.5 Hz), 7.72-7.83 (m, 2H), 7.60-7.70 (m, 2H),7.53 (d, 1H, J=8.6 Hz), 7.28-7.42 (m, 2H), 5.52 (s, 2H), 2.68 (t, 2H,J=7.4 Hz), 1.50-1.67 (m, 2H), 1.03-1.26 (m, 4H), and 0.67 ppm (t, 3H,J=7.0 Hz).

Step 3: Synthesis of5-(3-Pentyl-1-benzofuran-2-yl)-8-(1H-tetraazol-5-ylmethoxy)quinoline:The title compound was prepared as a yellow solid (0.044 g, 67%) from{[5-(3-pentyl-1-benzofuran-2-yl)-quinolin-8-yl]oxy}acetonitrileaccording to the procedure described in Step 3 of Example 1, mp 226-228°C. (dec.). Mass spectrum (+ESI, [M+H)⁺) m/z 414. ¹H NMR (500 MHz,DMSO-d₆): δ14.95-17.95 (brs, 1H), 9.01 (d, 1H, J=4.1 Hz), 8.35 (d, 1H,J=8.4 Hz), 7.81 (d, 1H, J=8.1 Hz), 7.71-7.77 (m, 2H), 7.60-7.65 (m, 2H),7.30-7.39 (m, 2H), 5.84 (s, 2H), 2.66 (t, 2H, J=7.6 Hz), 1.54-1.63 (m,2H), 1.06-1.18 (m, 4H), and 0.67 ppm (t, 3H, J=7.0 Hz). ElementalAnalysis for C₂₄H₂₃N₅O₂.HCl: Calculated: C, 64.07; H, 5.38; N, 15.57.Found: C, 63.97; H, 5.35; N, 15.47.

Example 6 Primary Screen for the PAI-1 Inhibition

Test compounds are dissolved in DMSO at a final concentration of 10 mM,then diluted 100× in physiologic buffer. The inhibitory assay isinitiated by the addition of the test compound (1-100 μM finalconcentration, maximum DMSO concentration of 0.2%) in a pH 6.6 buffercontaining 140 nM recombinant human plasminogen activator inhibitor-1(PAI-1; Molecular Innovations, Royal Oak, Mich.). Following a 1 hourincubation at room temperature, 70 nM of recombinant human tissueplasminogen activator (tPA) is added, and the combination of the testcompound, PAI-1 and tPA is incubated for an additional 30 minutes.Following the second incubation, Spectrozyme-tPA (American Diagnostica,Greenwich, Conn.), a chromogenic substrate for tPA, is added andabsorbance read at 405 nm at 0 and 60 minutes. Relative PAI-1 inhibitionis equal to the residual tPA activity in the presence of the testcompound and PAI-1. Control treatments include the complete inhibitionof tPA by PAI-1 at the molar ratio employed (2:1), and the absence ofany effect of the test compound on tPA alone.

Example 7 Assay for Determining IC₅₀ of Inhibition of PAI-1

This assay is based upon the non-SDS dissociable interaction between tPAand active PAI-1. Assay plates are initially coated with human tPA (10μg/ml). Test compounds are dissolved in DMSO at 10 mM, then diluted withphysiologic buffer (pH 7.5) to a final concentration of 1-50 μM. Testcompounds are incubated with human PAI-1 (50 ng/ml) for 15 minutes atroom temperature. The tPA-coated plate is washed with a solution of0.05% Tween 20 and 0.1% BSA, then the plate is blocked with a solutionof 3% BSA. An aliquot of the test compound/PAI-1 solution is then addedto the tPA-coated plate, incubated at room temperature for 1 hour, andwashed. Active PAI-1 bound to the plate is assessed by adding an aliquotof a 1:1000 dilution of the 33B8 monoclonal antibody against humanPAI-1, and incubating the plate at room temperature for 1 hour(Molecular Innovations, Royal Oak, Mich.). The plate is again washed,and a solution of goat anti-mouse IgG-alkaline phosphatase conjugate isadded at a 1:50,000 dilution in goat serum. The plate is incubated 30minutes at room temperature, washed, and a solution of alkalinephosphatase substrate is added. The plate is incubated 45 minutes atroom temperature, and color development is determined at OD_(405nm). Thequantitation of active PAI-1 bound to tPA at varying concentrations oftest compound is used to determine the IC₅₀. Results are analyzed usinga logarithmic best-fit equation. The assay sensitivity is 5 ng/ml ofhuman PAI-1 as determined from a standard curve ranging from 0-100ng/ml.

The compounds of the present invention inhibited Plasminogen ActivatorInhibitor-1 as summarized in Table 1.

TABLE 1 % Inhibition @ Compound IC₅₀ (μM) 25 μM 1 38.6^(b) 2 21.46^(a) 3— 20 4 — 55 5 8.1^(b) ^(a)The IC₅₀ was determined by the Antibody Assaydescribed above. ^(b)The IC₅₀ was determined by a modification of thePrimary Screen for PAI-1 Inhibition

Although the foregoing invention has been described in detail by way ofexample for purposes of clarity of understanding, it will be apparent tothe artisan that certain changes and modifications are comprehended bythe disclosure and can be practiced without undue experimentation withinthe scope of the appended claims, which are presented by way ofillustration not limitation.

All publications and patent documents cited above are herebyincorporated by reference in their entirety for all purposes to the sameextent as if each were so individually denoted.

1. A compound of the formula:

or a pharmaceutically acceptable salt or ester form thereof wherein: R,R₁, R₂, and R₃ are, independently, hydrogen, alkyl, cycloalkyl,alkanoyl, halo, hydroxy, aryl, perfluoroalkyl, alkoxy, amino,alkylamino, dialkylamino, or perfluoroalkoxy; R₄ is hydrogen, alkyl,perfluoroalkyl, alkenyl, alkynyl, arylalkenyl, arylalkynyl, aryl,—C(═O)R₅, —C(═S)R₅, —CH₂R₅, or —CH(OH)R₅; R₅ is hydrogen, C₁-C₈ alkyl,C₃-C₈ cycloalkyl, perfluoroalkyl, aryl, alkenyl, alkynyl, arylalkyl,arylalkenyl, arylalkynyl, alkylaryl, alkenylaryl, or alkynylaryl; R₆ ishydrogen, alkyl, aryl, or arylalkyl; x₁, x₂, x₃, x₄, x₅, x₆, x₇, and x₈are, independently, carbon or nitrogen, wherein at least one of x₁-x₈ isa nitrogen atom; n is an integer from 0-6; and A is COOH or an acidmimic or a pharmaceutically acceptable salt thereof.
 2. A compound ofthe formula:

wherein: R, R₁, R₂, and R₃ are, independently, hydrogen, alkyl,cycloalkyl, alkanoyl, halo, hydroxy, aryl, perfluoroalkyl, alkoxy,amino, alkylamino, dialkylamino, or perfluoroalkoxy; R₄ is hydrogen,alkyl, perfluoroalkyl, alkenyl, alkynyl, arylalkenyl, arylalkynyl, aryl,—C(═O)R₅, —C(═S)R₅, —CH₂R₅, or —CH(OH)R₅; R₅ is hydrogen, C₁-C₈ alkyl,C₃-C₈ cycloalkyl, perfluoroalkyl, aryl, alkenyl, alkynyl, arylalkyl,arylalkenyl, arylalkynyl, alkylaryl, alkenylaryl, or alkynylaryl; R₆ ishydrogen, alkyl, aryl, or arylalkyl; x₁, x₂, x₃, x₄, x₅, x₆, x₇, and x₈are, independently, carbon or nitrogen, wherein at least one of x₁-x₈ isa nitrogen atom; and A is an acid mimic or a pharmaceutically acceptablesalt thereof.
 3. The compound of claim 1 or 2, wherein A is tetrazole,tetronic acid, acyl tetronic acid, or a group having the formula:

wherein R₁₅ is C₁-C₆ alkyl, C₂-C₆ alkenyl, C₃-C₆ cycloalkyl, —CH₂—(C₃-C₆cycloalkyl), C₃-C₆ cycloalkenyl, —CH₂—(C₃-C₆ cycloalkenyl), optionallysubstituted aryl or heteroaryl groups or optionally substitutedaryl(C₁-C₆)alkyl or heteroaryl(C₁-C₆)alkyl, or a pharmaceuticallyacceptable salt thereof.
 4. The compound of claims 1 or 2, wherein A istetrazole or a pharmaceutically acceptable salt thereof.
 5. The compoundof claims 1 or 2, wherein R and R₁ are hydrogen or a pharmaceuticallyacceptable salt thereof.
 6. The compound of claims 1 or 2, wherein R₄ ishydrogen, alkyl, or aryl or a pharmaceutically acceptable salt thereof.7. The compound of claims 1 or 2, wherein R₂ and R₃ are, independently,hydrogen or perfluoroalkyl or a pharmaceutically acceptable saltthereof.
 8. The compound of claims 1 or 2, wherein R₄ is perfluoroalkyl,alkenyl, arylalkenyl, aryl, —C(—O)R₅, —C(═S)R₅, —CH₃R₅, or —CH₂(OH)R₅,or a pharmaceutically acceptable salt thereof.
 9. The compound of claim2 that is 6-(1-benzofuran-2-yl)-2-(trifluoromethyl)quinolin-4-yl1H-tetraazol-5-ylmethyl ether or a pharmaceutically acceptable salt orester form thereof.
 10. The compound of claim 2 that is6-(3-Pentyl-1-benzofuran-2-yl)-4-(1H-tetraazol-5-ylmethoxy)-2-(trifluoromethyl)-quinolineor a pharmaceutically acceptable salt or ester form thereof.
 11. Thecompound of claim 2 that is5-(1-Benzofuran-2-yl)-8-(1H-tetraazol-5-ylmethoxy)quinoline or apharmaceutically acceptable salt or ester form thereof.
 12. The compoundof claim 2 that is 5-(3-Phenyl-1-benzofuran-2-yl)-8-quinolinyl1H-tetraazol-5-ylmethyl ether or a pharmaceutically acceptable salt orester form thereof.
 13. The compound of claim 2 that is5-(3-Pentyl-1-benzofuran-2-yl)-8-(1H-tetraazol-5-ylmethoxy)quinoline ora pharmaceutically acceptable salt or ester form thereof.
 14. A methodof inhibiting PAI-1 activity comprising administering to a subject inneed thereof a therapeutically effective amount of the compound ofclaims 1 or
 2. 15. The method of claim 14, wherein the therapeuticallyeffective amount is from 25 mg/kg/day to 200 mg/kg/day.
 16. A method fortreating a PAI-1 related disorder in a subject, the method comprisingidentifying a subject having a PAI-1 related disorder and administeringto a subject in need thereof a therapeutically effective amount of thecompound of claims 1 or
 2. 17. The method of claim 16, wherein thetherapeutically effective amount is from 25 mg/kg/day to 200 mg/kg/day.18. The method of claim 16, wherein the PAI-1 related disorder isimpairment of the fibrinolytic system.
 19. The method of claim 16,wherein the PAI-1 related disorder is thrombosis, atrial fibrillation,pulmonary fibrosis, thromboembolic complication of surgery,cardiovascular disease, myocardial ischemia, stroke, atheroscleroticplaque formation, chronic obstructive pulmonary disease, renal fibrosis,polycystic ovary syndrome, diabetes, Alzheimer's disease, or cancer. 20.The method of claim 19, wherein the thrombosis is selected from thegroup consisting of venous thrombosis, arterial thrombosis, cerebralthrombosis, and deep vein thrombosis.
 21. The method of claim 19,wherein the PAI-1 related disorder is cardiovascular disease caused bynoninsulin dependent diabetes mellitus in a mammal.
 22. A pharmaceuticalcomposition comprising a therapeutically effective amount of a compoundof claims 1 or 2, or a pharmaceutically acceptable salt or ester formthereof, and a pharmaceutically acceptable excipient or carrier.
 23. Amethod for treating thrombosis, atrial fibrillation, pulmonary fibrosis,thromboembolic complication of surgery, stroke, myocardial ischemia,atherosclerotic plaque formation, chronic obstructive pulmonary disease,or renal fibrosis comprising administering to a subject in need thereofa therapeutically effective amount of a compound of formula 1:

or a pharmaceutically acceptable salt or ester form thereof wherein: R,R₁, R₂, and R₃ are, independently, hydrogen, alkyl, cycloalkyl,alkanoyl, halo, hydroxy, aryl, perfluoroalkyl, alkoxy, amino,alkylamino, dialkylamino, or perfluoroalkoxy; R₄ is hydrogen, alkyl,perfluoroalkyl, alkenyl, alkynyl, arylalkenyl, arylalkynyl, aryl,—C(═O)R₅, —C(═S)R₅, —CH₂R₅, or —CH(OH)R₅. R₅ is hydrogen, C₁-C₈ alkyl,C₃-C₈ cycloalkyl, perfluoroalkyl, aryl, alkenyl, alkynyl, arylalkyl,arylalkenyl, arylalkynyl, alkylaryl, alkenylaryl, or alkynylaryl; R₆ ishydrogen, alkyl, aryl, or arylalkyl; x₁, x₂, x₃, x₄, x₅, x₆, x₇, and x₈are, independently, carbon or nitrogen, wherein at least one of x₁-x₈ isa nitrogen atom; n is an integer from 1-6; and A is COOH or an acidmimic.
 24. A method for treating a PAI-1 related disorder in a subject,the method comprising identifying a subject having a PAI-1 relateddisorder and administering to a subject in need thereof atherapeutically effective amount of the compound of claim
 2. 25. Themethod of claim 24, wherein the therapeutically effective amount is from25 mg/kg/day to 200 mg/kg/day.
 26. The method of claim 24, wherein thePAI-1 related disorder is impairment of the fibrinolytic system.
 27. Themethod of claim 24, wherein the PAI-1 related disorder is thrombosis,atrial fibrillation, pulmonary fibrosis, thromboembolic complication ofsurgery, cardiovascular disease, myocardial ischemia, stroke,atherosclerotic plaque formation, chronic obstructive pulmonary disease,renal fibrosis, polycystic ovary syndrome, diabetes, Alzheimer'sdisease, or cancer.
 28. The method of claim 27, wherein the thrombosisis selected from the group consisting of venous thrombosis, arterialthrombosis, cerebral thrombosis, and deep vein thrombosis.
 29. Themethod of claim 24, wherein the PAI-1 related disorder is cardiovasculardisease caused by noninsulin dependent diabetes mellitus in a mammal.30. A method for treating thrombosis, atrial fibrillation, pulmonaryfibrosis, thromboembolic complication of surgery, stroke, myocardialischemia, atherosclerotic plaque formation, chronic obstructivepulmonary disease, or renal fibrosis comprising administering to asubject in need thereof a therapeutically effective amount of a compoundof formula 2:

wherein: R, R₁, R₂, and R₃ are, independently, hydrogen, alkyl,cycloalkyl, alkanoyl, halo, hydroxy, aryl, perfluoroalkyl, alkoxy,amino, alkylamino, dialkylamino, or perfluoroalkoxy; R₄ is hydrogen,alkyl, perfluoroalkyl, alkenyl, alkynyl, arylalkenyl, arylalkynyl, aryl,—C(═O)R₅, —C(═S)R₅, —CH₂R₅, or —CH(OH)R₅. R₅ is hydrogen, C₁-C₈ alkyl,C₃-C₈ cycloalkyl, perfluoroalkyl, aryl, alkenyl, alkynyl, arylalkyl,arylalkenyl, arylalkynyl, alkylaryl, alkenylaryl, or alkynylaryl; R₆ ishydrogen, alkyl, aryl, or arylalkyl; x₁, x₂, x₃, x₄, x₅, x₆, x₇, and x₈are, independently, carbon or nitrogen, wherein at least one of x₁-x₈ isa nitrogen atom; and A is an acid mimic.